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Sample records for extracellular matrix affected

  1. Process-induced extracellular matrix alterations affect the mechanisms of soft tissue repair and regeneration

    PubMed Central

    Xu, Hui; Sandor, Maryellen; Lombardi, Jared

    2013-01-01

    Extracellular matrices derived from animal tissues for human tissue repairs are processed by various methods of physical, chemical, or enzymatic decellularization, viral inactivation, and terminal sterilization. The mechanisms of action in tissue repair vary among bioscaffolds and are suggested to be associated with process-induced extracellular matrix modifications. We compared three non-cross-linked, commercially available extracellular matrix scaffolds (Strattice, Veritas, and XenMatrix), and correlated extracellular matrix alterations to in vivo biological responses upon implantation in non-human primates. Structural evaluation showed significant differences in retaining native tissue extracellular matrix histology and ultrastructural features among bioscaffolds. Tissue processing may cause both the condensation of collagen fibers and fragmentation or separation of collagen bundles. Calorimetric analysis showed significant differences in the stability of bioscaffolds. The intrinsic denaturation temperature was measured to be 51°C, 38°C, and 44°C for Strattice, Veritas, and XenMatrix, respectively, demonstrating more extracellular matrix modifications in the Veritas and XenMatrix scaffolds. Consequently, the susceptibility to collagenase degradation was increased in Veritas and XenMatrix when compared to their respective source tissues. Using a non-human primate model, three bioscaffolds were found to elicit different biological responses, have distinct mechanisms of action, and yield various outcomes of tissue repair. Strattice permitted cell repopulation and was remodeled over 6 months. Veritas was unstable at body temperature, resulting in rapid absorption with moderate inflammation. XenMatrix caused severe inflammation and sustained immune reactions. This study demonstrates that extracellular matrix alterations significantly affect biological responses in soft tissue repair and regeneration. The data offer useful insights into the rational design of

  2. Static stretch affects neural stem cell differentiation in an extracellular matrix-dependent manner

    NASA Astrophysics Data System (ADS)

    Arulmoli, Janahan; Pathak, Medha M.; McDonnell, Lisa P.; Nourse, Jamison L.; Tombola, Francesco; Earthman, James C.; Flanagan, Lisa A.

    2015-02-01

    Neural stem and progenitor cell (NSPC) fate is strongly influenced by mechanotransduction as modulation of substrate stiffness affects lineage choice. Other types of mechanical stimuli, such as stretch (tensile strain), occur during CNS development and trauma, but their consequences for NSPC differentiation have not been reported. We delivered a 10% static equibiaxial stretch to NSPCs and examined effects on differentiation. We found static stretch specifically impacts NSPC differentiation into oligodendrocytes, but not neurons or astrocytes, and this effect is dependent on particular extracellular matrix (ECM)-integrin linkages. Generation of oligodendrocytes from NSPCs was reduced on laminin, an outcome likely mediated by the α6 laminin-binding integrin, whereas similar effects were not observed for NSPCs on fibronectin. Our data demonstrate a direct role for tensile strain in dictating the lineage choice of NSPCs and indicate the dependence of this phenomenon on specific substrate materials, which should be taken into account for the design of biomaterials for NSPC transplantation.

  3. Ameloblastin, an Extracellular Matrix Protein, Affects Long Bone Growth and Mineralization.

    PubMed

    Lu, Xuanyu; Fukumoto, Satoshi; Yamada, Yoshihiko; Evans, Carla A; Diekwisch, Thomas Gh; Luan, Xianghong

    2016-06-01

    Matrix molecules such as the enamel-related calcium-binding phosphoprotein ameloblastin (AMBN) are expressed in multiple tissues, including teeth, bones, and cartilage. Here we have asked whether AMBN is of functional importance for timely long bone development and, if so, how it exerts its function related to osteogenesis. Adolescent AMBN-deficient mice (AMBN(Δ5-6) ) suffered from a 33% to 38% reduction in femur length and an 8.4% shorter trunk spinal column when compared with WT controls, whereas there was no difference between adult animals. On a cellular level, AMBN truncation resulted in a shortened growth plate and a 41% to 49% reduction in the number of proliferating tibia chondrocytes and osteoblasts. Bone marrow stromal cells (BMSCs) isolated from AMBN mutant mice displayed defects in proliferation and differentiation potential as well as cytoskeleton organization. Osteogenesis-related growth factors, such as insulin-like growth factor 1 (IGF1) and BMP7, were also significantly (46% to 73%) reduced in AMBN-deficient BMSCs. Addition of exogenous AMBN restored cytoskeleton structures in AMBN mutant BMSCs and resulted in a dramatic 400% to 600% increase in BMP2, BMP7, and Col1A expression. Block of RhoA diminished the effect of AMBN on osteogenic growth factor and matrix protein gene expression. Addition of exogenous BMP7 and IGF1 rescued the proliferation and differentiation potential of AMBN-deficient BMSCs. Confirming the effects of AMBN on long bone growth, back-crossing of mutant mice with full-length AMBN overexpressors resulted in a complete rescue of AMBN(Δ5-6) bone defects. Together, these data indicate that AMBN affects extracellular matrix production and cell adhesion properties in the long bone growth plate, resulting in altered cytoskeletal dynamics, increased osteogenesis-related gene expression, as well as osteoblast and chondrocyte proliferation. We propose that AMBN facilitates rapid long bone growth and an important growth spurt during the

  4. Tendon Functional Extracellular Matrix

    PubMed Central

    Screen, H.R.C.; Birk, D.E.; Kadler, K.E.; Ramirez, F; Young, M.F.

    2015-01-01

    This article is one of a series, summarising views expressed at the Orthopaedic Research Society New Frontiers in Tendon Research Conference. This particular article reviews the three workshops held under the “Functional Extracellular Matrix” stream. The workshops focused on the roles of the tendon extracellular matrix, such as performing the mechanical functions of tendon, creating the local cell environment and providing cellular cues. Tendon is a complex network of matrix and cells, and its biological functions are influenced by widely-varying extrinsic and intrinsic factors such as age, nutrition, exercise levels and biomechanics. Consequently, tendon adapts dynamically during development, ageing and injury. The workshop discussions identified research directions associated with understanding cell-matrix interactions to be of prime importance for developing novel strategies to target tendon healing or repair. PMID:25640030

  5. Extracellular matrix and wound healing.

    PubMed

    Maquart, F X; Monboisse, J C

    2014-04-01

    Extracellular matrix has been known for a long time as an architectural support for the tissues. Many recent data, however, have shown that extracellular matrix macromolecules (collagens, elastin, glycosaminoglycans, proteoglycans and connective tissue glycoproteins) are able to regulate many important cell functions, such as proliferation, migration, protein synthesis or degradation, apoptosis, etc., making them able to play an important role in the wound repair process. Not only the intact macromolecules but some of their specific domains, that we called "Matrikines", are also able to regulate many cell activities. In this article, we will summarize main findings showing the effects of extracellular matrix macromolecules and matrikines on connective tissue and epithelial cells, particularly in skin, and their potential implication in the wound healing process. These examples show that extracellular matrix macromolecules or some of their specific domains may play a major role in wound healing. Better knowledge of these interactions may suggest new therapeutic targets in wound healing defects. PMID:24650524

  6. Involvement of extracellular matrix constituents in breast cancer

    SciTech Connect

    Lochter, Andre; Bissell, Mina J

    1995-06-01

    It has recently been established that the extracellular matrix is required for normal functional differentiation of mammary epithelia not only in culture, but also in vivo. The mechanisms by which extracellular matrix affects differentiation, as well as the nature of extracellular matrix constituents which have major impacts on mammary gland function, have only now begun to be dissected. The intricate variety of extracellular matrix-mediated events and the remarkable degree of plasticity of extracellular matrix structure and composition at virtually all times during ontogeny, make such studies difficult. Similarly, during carcinogenesis, the extracellular matrix undergoes gross alterations, the consequences of which are not yet precisely understood. Nevertheless, an increasing amount of data suggests that the extracellular matrix and extracellular matrix-receptors might participate in the control of most, if not all, of the successive stages of breast tumors, from appearance to progression and metastasis.

  7. Extracellular Matrix Abnormalities in Schizophrenia

    PubMed Central

    Berretta, Sabina

    2011-01-01

    Emerging evidence points to the involvement of the brain extracellular matrix (ECM) in the pathophysiology of schizophrenia (SZ). Abnormalities affecting several ECM components, including Reelin and chondroitin sulfate proteoglycans (CSPGs), have been described in subjects with this disease. Solid evidence supports the involvement of Reelin, an ECM glycoprotein involved in corticogenesis, synaptic functions and glutamate NMDA receptor regulation, expressed prevalently in distinct populations of GABAergic neurons, which secrete it into the ECM. Marked changes of Reelin expression in SZ have typically been reported in association with GABA-related abnormalities in subjects with SZ and bipolar disorder. Recent findings from our group point to substantial abnormalities affecting CSPGs, a main ECM component, in the amygdala and entorhinal cortex of subjects with schizophrenia, but not bipolar disorder. Striking increases of glial cells expressing CSPGs were accompanied by reductions of perineuronal nets, CSPG- and Reelin-enriched ECM aggregates enveloping distinct neuronal populations. CSPGs developmental and adult functions, including neuronal migration, axon guidance, synaptic and neurotransmission regulation are highly relevant to the pathophysiology of SZ. Together with reports of anomalies affecting several other ECM components, these findings point to the ECM as a key component of the pathology of SZ. We propose that ECM abnormalities may contribute to several aspects of the pathophysiology of this disease, including disrupted connectivity and neuronal migration, synaptic anomalies and altered GABAergic, glutamatergic and dopaminergic neurotransmission. PMID:21856318

  8. Cleavage of extracellular matrix in periodontitis: gingipains differentially affect cell adhesion activities of fibronectin and tenascin-C

    PubMed Central

    Ruggiero, Sabrina; Cosgarea, Raluca; Potempa, Jan; Potempa, Barbara; Eick, Sigrun; Chiquet, Matthias

    2014-01-01

    Gingipains are cysteine proteases that represent major virulence factors of the periodontopathogenic bacterium Porphyromonas gingivalis. Gingipains are reported to degrade extracellular matrix (ECM) of periodontal tissues, leading to tissue destruction and apoptosis. The exact mechanism is not known, however. Fibronectin and tenascin-C are pericellular ECM glycoproteins present in periodontal tissues. Whereas fibronectin mediates fibroblast adhesion, tenascin-C binds to fibronectin and inhibits its cell-spreading activity. Using purified proteins in vitro, we asked whether fibronectin and tenascin-C are cleaved by gingipains at clinically relevant concentrations, and how fragmentation by the bacterial proteases affects their biological activity in cell adhesion. Fibronectin was cleaved into distinct fragments by all three gingipains; however, only arginine-specific HRgpA and RgpB but not lysine-specific Kgp destroyed its cell-spreading activity. This result was confirmed with recombinant cell-binding domain of fibronectin. Of the two major tenascin-C splice variants, the large but not the small was a substrate for gingipains, indicating that cleavage occurred primarily in the alternatively spliced domain. Surprisingly, cleavage of large tenascin-C variant by all three gingipains generated fragments with increased anti-adhesive activity towards intact fibronectin. Fibronectin and tenascin-C fragments were detected in gingival crevicular fluid of a subset of periodontitis patients. We conclude that cleavage by gingipains directly affects the biological activity of both fibronectin and tenascin-C in a manner that might lead to increased cell detachment and loss during periodontal disease. PMID:23313574

  9. Cleavage of extracellular matrix in periodontitis: gingipains differentially affect cell adhesion activities of fibronectin and tenascin-C.

    PubMed

    Ruggiero, Sabrina; Cosgarea, Raluca; Potempa, Jan; Potempa, Barbara; Eick, Sigrun; Chiquet, Matthias

    2013-04-01

    Gingipains are cysteine proteases that represent major virulence factors of the periodontopathogenic bacterium Porphyromonas gingivalis. Gingipains are reported to degrade extracellular matrix (ECM) of periodontal tissues, leading to tissue destruction and apoptosis. The exact mechanism is not known, however. Fibronectin and tenascin-C are pericellular ECM glycoproteins present in periodontal tissues. Whereas fibronectin mediates fibroblast adhesion, tenascin-C binds to fibronectin and inhibits its cell-spreading activity. Using purified proteins in vitro, we asked whether fibronectin and tenascin-C are cleaved by gingipains at clinically relevant concentrations, and how fragmentation by the bacterial proteases affects their biological activity in cell adhesion. Fibronectin was cleaved into distinct fragments by all three gingipains; however, only arginine-specific HRgpA and RgpB but not lysine-specific Kgp destroyed its cell-spreading activity. This result was confirmed with recombinant cell-binding domain of fibronectin. Of the two major tenascin-C splice variants, the large but not the small was a substrate for gingipains, indicating that cleavage occurred primarily in the alternatively spliced domain. Surprisingly, cleavage of large tenascin-C variant by all three gingipains generated fragments with increased anti-adhesive activity towards intact fibronectin. Fibronectin and tenascin-C fragments were detected in gingival crevicular fluid of a subset of periodontitis patients. We conclude that cleavage by gingipains directly affects the biological activity of both fibronectin and tenascin-C in a manner that might lead to increased cell detachment and loss during periodontal disease. PMID:23313574

  10. Contact guidance induced organization of extracellular matrix.

    PubMed

    Manwaring, Michael E; Walsh, Jennifer F; Tresco, Patrick A

    2004-08-01

    The scarring response following injury to the central nervous system disrupts the anatomical organization of nervous tissue posing a barrier to the regeneration of axons. In the present study, using materials with nanometer level surface features we examined whether matrix organization could be controlled by engineering meningeal cell asymmetry. Following 5 days in culture, the organization of meningeal cells along with their cytoskeletal elements and extracellular matrix proteins was evaluated. Meningeal cell morphology was markedly affected by nanometer level substrate topography. Cell alignment increased with increasing surface roughness. In addition, linear arrays of extracellular matrix were expressed that appeared related to cellular orientation. When cultured on substrates with topographical features of less than 10 nm neither cells nor their extracellular matrix showed organizational asymmetry. However, as oriented surface roughness increased, cellular and matrix asymmetrical organization became more pronounced reaching a threshold at 345 nm. These results suggest that biomaterial surface topography or other methods of altering the orientation of cells may be used to engineer orientation into the secreted extracellular matrix and as such may be a potential strategy for developing organized cell-derived matrix as a bridging material for nerve repair or other regenerative applications. PMID:15020137

  11. Mechanotransduction and extracellular matrix homeostasis

    PubMed Central

    Humphrey, Jay D.; Dufresne, Eric R.; Schwartz, Martin A.

    2015-01-01

    Preface Soft connective tissues at steady state are yet dynamic; resident cells continually read environmental cues and respond to promote homeostasis, including maintenance of the mechanical properties of the extracellular matrix that are fundamental to cellular and tissue health. The mechanosensing process involves assessment of the mechanics of the matrix by the cells through integrins and the actomyosin cytoskeleton, and is followed by a mechano-regulation process that includes the deposition, rearrangement, or removal of matrix to maintain overall form and function. Progress toward understanding the molecular, cellular, and tissue scale effects that promote mechanical homeostasis has helped identify key questions for future research. PMID:25355505

  12. Extracellular matrix in ovarian follicles.

    PubMed

    Rodgers, R J; Irving-Rodgers, H F; van Wezel, I L

    2000-05-25

    A lot is known about the control of the development of ovarian follicles by growth factors and hormones, but less is known about the roles of extracellular matrix in the control of follicular growth and development. In this review we focus on the specialized extracellular matrix of the basal laminas that are present in ovarian follicles. These include the follicular basal lamina itself, the Call-Exner bodies of the membrana granulosa, the subendothelial and arteriole smooth muscle basal laminas in the theca, and the basal lamina-like material of the thecal matrix. We discuss the evidence that during follicle development the follicular basal lamina changes in composition, that many of its components are produced by the granulosa cells, and that the follicular basal laminas of different follicles have different ultrastructural appearances, linked to the shape of the aligning granulosa cells. All these studies suggest that the follicular basal lamina is extremely dynamic during follicular development. PMID:10963877

  13. Low-Intensity Pulsed Ultrasound Affects Chondrocyte Extracellular Matrix Production via an Integrin-Mediated p38 MAPK Signaling Pathway.

    PubMed

    Xia, Peng; Ren, Shasha; Lin, Qiang; Cheng, Kai; Shen, Shihao; Gao, Mingxia; Li, Xueping

    2015-06-01

    Although low-intensity pulsed ultrasound (LIPUS) regulates p38 mitogen-activated protein kinase (MAPK) and promotes cartilage repair in osteoarthritis, the role of integrin-mediated p38 MAPK in the effect of LIPUS on extracellular matrix (ECM) production of normal and OA chondrocytes remains unknown. The aim of this study was to investigate whether LIPUS affects ECM production in normal and OA rabbit chondrocytes through an integrin-p38 signaling pathway. A rabbit model of OA was established by anterior cruciate ligament transection, and chondrocytes were isolated from normal or OA cartilage and cultured in vitro. Chondrocytes were treated with LIPUS and then pre-incubated with the integrin inhibitor GRGDSP or the p38 inhibitor SB203580. Expression of type II collagen, MMP-13, integrin β1, p38 and phosphorylated p38 was assessed by Western blot analysis. We found that type II collagen and integrin β1 were upregulated (p < 0.05), whereas MMP-13 was downregulated (p < 0.05) in normal and OA chondrocytes. Furthermore, phosphorylated p38 was upregulated (p < 0.05) in normal chondrocytes, but downregulated (p < 0.05) in OA chondrocytes after LIPUS stimulation. Pre-incubation of chondrocytes with the integrin inhibitor disrupted the effects of LIPUS on normal and OA chondrocytes. Pre-incubation of chrondocytes with the p38 inhibitor reduced the effects of LIPUS on normal chondrocytes, but had no impact on OA chondrocytes. Our findings suggest that the integrin-p38 MAPK signaling pathway plays an important role in LIPUS-mediated ECM production in chondrocytes. PMID:25736607

  14. Parenteral and enteral feeding in preterm piglets differently affects extracellular matrix proteins, enterocyte proliferation and apoptosis in the small intestine.

    PubMed

    Oste, Marijke; De Vos, Maartje; Van Haver, Els; Van Brantegem, Leen; Thymann, Thomas; Sangild, Per; Weyns, Andre; Van Ginneken, Chris

    2010-10-01

    The preterm intestine is immature and responds differently to total parenteral nutrition (TPN) and enteral nutrition, compared with the term intestine. We hypothesised that in preterms, diet composition and feeding route affect mucosal morphology, enterocyte mitosis and apoptosis, and the distribution of laminin-1, fibronectin and collagen IV (extracellular matrix proteins (ECMP)). Preterm piglets (93.5 % of gestation) were delivered via caesarean section and birth weight-matched allocated to one of the four experimental groups: the piglets were either euthanised immediately after delivery, after 3 d of TPN or after 2 d enteral feeding with colostrum or milk formula, following 3 d of TPN. We combined immunohistochemistry, image analysis and stereological measurements to describe the intestinal mucosal layer. No significant changes occurred after 3 d of TPN. Feeding colostrum or milk replacer for 2 d after TPN was associated with an increased crypt depth. Only enteral feeding with colostrum resulted in an increased villus height and mitotic index. Neither TPN nor enteral feeding changed the distribution pattern of ECMP or the occurrence of bifid crypts. The immature distribution pattern of ECMP in TPN-fed piglets, coupled with unchanged enterocyte mitosis and apoptosis indices, illustrates that feeding preterm pigs 3 d TPN does not lead to mucosal atrophy. Despite the invariable distribution of ECMP, colostrum was associated with crypt hyperplasia resulting in an increased villus height. These data illustrate that some mechanisms regulating cell turnover are immature in preterms and may in part explain the abnormal gut responses to TPN and enteral feeding in prematurely born pigs. PMID:20887647

  15. Brain Extracellular Matrix in Neurodegeneration

    PubMed Central

    Bonneh-Barkay, Dafna; Wiley, Clayton A.

    2009-01-01

    The role of extracellular matrix (ECM) in neurological development, function and degeneration has evolved from a simplistic physical adhesion to a system of intricate cellular signaling. While most cells require ECM adhesion to survive, it is now clear that differentiated function is intimately dependent upon cellular interaction with the ECM. Therefore, it is not surprising that the ECM is increasingly found to be involved in the enigmatic process of neurodegeneration. Descriptive studies of human neurodegenerative disorders and experimental studies of animal models of neurodegeneration have begun to define potential mechanisms of ECM disruption that can lead to synaptic and neuronal loss. PMID:18662234

  16. LTBP2 mutations cause Weill-Marchesani and Weill-Marchesani-like syndrome and affect disruptions in the extracellular matrix.

    PubMed

    Haji-Seyed-Javadi, Ramona; Jelodari-Mamaghani, Sahar; Paylakhi, Seyed Hassan; Yazdani, Shahin; Nilforushan, Naveed; Fan, Jian-Bing; Klotzle, Brandy; Mahmoudi, Mohammad Jafar; Ebrahimian, Mohammad Jafar; Chelich, Noori; Taghiabadi, Ehsan; Kamyab, Kambiz; Boileau, Catherine; Paisan-Ruiz, Coro; Ronaghi, Mostafa; Elahi, Elahe

    2012-08-01

    Latent transforming growth factor (TGF) beta-binding protein 2 (LTBP2) is an extracellular matrix (ECM) protein that associates with fibrillin-1 containing microfibrils. Various factors prompted considering LTBP2 in the etiology of isolated ectopia lentis and associated conditions such as Weill-Marchesani syndrome (WMS) and Marfan syndrome (MFS). LTBP2 was screened in 30 unrelated Iranian patients. Mutations were found only in one WMS proband and one MFS proband. Homozygous c.3529G>A (p.Val1177Met) was shown to cause autosomal recessive WMS or WM-like syndrome by several approaches, including homozygosity mapping. Light, fluorescent, and electron microscopy evidenced disruptions of the microfibrillar network in the ECM of the proband's skin. In conjunction with recent findings regarding other ECM proteins, the results presented strongly support the contention that anomalies in WMS patients are due to disruptions in the ECM. Heterozygous c.1642C >T (p.Arg548*) possibly contributed to MFS-related phenotypes, including ocular manifestations, mitral valve prolapse, and pectus excavatum, but was not cause of MFS. PMID:22539340

  17. The Evolution of Extracellular Matrix

    PubMed Central

    Özbek, Suat; Balasubramanian, Prakash G.; Chiquet-Ehrismann, Ruth; Tucker, Richard P.

    2010-01-01

    We present a perspective on the molecular evolution of the extracellular matrix (ECM) in metazoa that draws on research publications and data from sequenced genomes and expressed sequence tag libraries. ECM components do not function in isolation, and the biological ECM system or “adhesome” also depends on posttranslational processing enzymes, cell surface receptors, and extracellular proteases. We focus principally on the adhesome of internal tissues and discuss its origins at the dawn of the metazoa and the expansion of complexity that occurred in the chordate lineage. The analyses demonstrate very high conservation of a core adhesome that apparently evolved in a major wave of innovation in conjunction with the origin of metazoa. Integrin, CD36, and certain domains predate the metazoa, and some ECM-related proteins are identified in choanoflagellates as predicted sequences. Modern deuterostomes and vertebrates have many novelties and elaborations of ECM as a result of domain shuffling, domain innovations and gene family expansions. Knowledge of the evolution of metazoan ECM is important for understanding how it is built as a system, its roles in normal tissues and disease processes, and has relevance for tissue engineering, the development of artificial organs, and the goals of synthetic biology. PMID:21160071

  18. Extracellular matrix as target for antitumor therapy

    PubMed Central

    Harisi, Revekka; Jeney, Andras

    2015-01-01

    The aim of the present review is to survey the accumulated knowledge on the extracellular matrix (ECM) of tumors referring to its putative utility as therapeutic target. Following the traditional observation on the extensive morphological alteration in the tumor-affected tissue, the well-documented aberrant cellular regulation indicated that ECM components have an active role in tumor progression. However, due to the diverse functions and variable expression of proteoglycans, matrix proteins, and integrins, it is rather difficult to identify a comprehensive therapeutic target among ECM components. At present, the elevated level of heparanase and the prominent expression of αvβ5 integrin are considered as promising therapeutic targets. The inhibition of glycosaminoglycan offers another promising approach in the treatment of those tumors which are stimulated by proteoglycans. It can be ascertained that a selective ECM inhibitor would be a great asset to control metastasis driven by ECM-mediated signaling. PMID:26089687

  19. Stretching the boundaries of extracellular matrix research.

    PubMed

    Hynes, Richard O

    2014-12-01

    Extracellular matrix (ECM) proteins constitute >1% of the proteome and interact with many modifiers and growth factors to affect most aspects of cellular behaviour during development and normal physiology, as well as in diseases such as fibroses, cancer and many genetic disorders. In addition to biochemical signals provided to cells by ECM proteins, important cell–ECM interactions involve bidirectional mechanotransduction influences, which are dependent on the physical structure and organization of the ECM. These are beginning to be understood using twenty-first-century approaches, including biophysics, nanotechnology, biological engineering and modern microscopy. Articles in this issue of Nature Reviews Molecular Cell Biology review progress in our understanding of the ECM. PMID:25574535

  20. Molecular Adhesion between Cartilage Extracellular Matrix Macromolecules

    PubMed Central

    2015-01-01

    In this study, we investigated the molecular adhesion between the major constituents of cartilage extracellular matrix, namely, the highly negatively charged proteoglycan aggrecan and the type II/IX/XI fibrillar collagen network, in simulated physiological conditions. Colloidal force spectroscopy was applied to measure the maximum adhesion force and total adhesion energy between aggrecan end-attached spherical tips (end radius R ≈ 2.5 μm) and trypsin-treated cartilage disks with undamaged collagen networks. Studies were carried out in various aqueous solutions to reveal the physical factors that govern aggrecan–collagen adhesion. Increasing both ionic strength and [Ca2+] significantly increased adhesion, highlighting the importance of electrostatic repulsion and Ca2+-mediated ion bridging effects. In addition, we probed how partial enzymatic degradation of the collagen network, which simulates osteoarthritic conditions, affects the aggrecan–collagen interactions. Interestingly, we found a significant increase in aggrecan–collagen adhesion even when there were no detectable changes at the macro- or microscales. It is hypothesized that the aggrecan–collagen adhesion, together with aggrecan–aggrecan self-adhesion, works synergistically to determine the local molecular deformability and energy dissipation of the cartilage matrix, in turn, affecting its macroscopic tissue properties. PMID:24491174

  1. Proteases decode the extracellular matrix cryptome.

    PubMed

    Ricard-Blum, Sylvie; Vallet, Sylvain D

    2016-03-01

    The extracellular matrix is comprised of 1100 core-matrisome and matrisome-associated proteins and of glycosaminoglycans. This structural scaffold contributes to the organization and mechanical properties of tissues and modulates cell behavior. The extracellular matrix is dynamic and undergoes constant remodeling, which leads to diseases if uncontrolled. Bioactive fragments, called matricryptins, are released from the extracellular proteins by limited proteolysis and have biological activities on their own. They regulate numerous physiological and pathological processes such as angiogenesis, cancer, diabetes, wound healing, fibrosis and infectious diseases and either improve or worsen the course of diseases depending on the matricryptins and on the molecular and biological contexts. Several protease families release matricryptins from core-matrisome and matrisome-associated proteins both in vitro and in vivo. The major proteases, which decrypt the extracellular matrix, are zinc metalloproteinases of the metzincin superfamily (matrixins, adamalysins and astacins), cysteine proteinases and serine proteases. Some matricryptins act as enzyme inhibitors, further connecting protease and matricryptin fates and providing intricate regulation of major physiopathological processes such as angiogenesis and tumorigenesis. They strengthen the role of the extracellular matrix as a key player in tissue failure and core-matrisome and matrisome-associated proteins as important therapeutic targets. PMID:26382969

  2. Focused Low-intensity Pulsed Ultrasound Affects Extracellular Matrix Degradation via Decreasing Chondrocyte Apoptosis and Inflammatory Mediators in a Surgically Induced Osteoarthritic Rabbit Model.

    PubMed

    Jia, Lang; Chen, Jinyun; Wang, Yan; Zhang, Yu; Chen, Wenzhi

    2016-01-01

    We investigated whether focused low-intensity pulsed ultrasound (FLIPUS) affects extracellular matrix (ECM) production in osteoarthritic (OA) rabbits by decreasing chondrocyte apoptosis and pro-inflammatory mediators. An OA model using New Zealand White rabbits (N = 30) and 30 normal rabbits were randomized into three groups (2-, 4- and 8-wk groups; n = 10 knees each). A knee from each rabbit was randomly selected to receive FLIPUS and the other knee received a sham treatment as a control. Another 30 normal rabbits were blank controls. We measured ECM degradation, joint effusion volume and levels of prostaglandin E2 and nitric oxide. Also, ratios of chondrocyte proliferation and apoptosis were calculated. Compared with sham stimulation, FLIPUS attenuated release of type II collagen and proteoglycans and reduced chondrocyte apoptosis as well as total joint effusion volume and significantly alleviated OA-induced accretion of prostaglandin E2 and nitric oxide in the synovial fluid. FLIPUS application promoted ECM production in OA through down regulation inflammatory mediators, joint effusion volume and chondrocyte apoptosis. PMID:26403700

  3. Extracellular matrix component signaling in cancer.

    PubMed

    Multhaupt, Hinke A B; Leitinger, Birgit; Gullberg, Donald; Couchman, John R

    2016-02-01

    Cell responses to the extracellular matrix depend on specific signaling events. These are important from early development, through differentiation and tissue homeostasis, immune surveillance, and disease pathogenesis. Signaling not only regulates cell adhesion cytoskeletal organization and motility but also provides survival and proliferation cues. The major classes of cell surface receptors for matrix macromolecules are the integrins, discoidin domain receptors, and transmembrane proteoglycans such as syndecans and CD44. Cells respond not only to specific ligands, such as collagen, fibronectin, or basement membrane glycoproteins, but also in terms of matrix rigidity. This can regulate the release and subsequent biological activity of matrix-bound growth factors, for example, transforming growth factor-β. In the environment of tumors, there may be changes in cell populations and their receptor profiles as well as matrix constitution and protein cross-linking. Here we summarize roles of the three major matrix receptor types, with emphasis on how they function in tumor progression. PMID:26519775

  4. Nanomechanics of the Cartilage Extracellular Matrix

    NASA Astrophysics Data System (ADS)

    Han, Lin; Grodzinsky, Alan J.; Ortiz, Christine

    2011-08-01

    Cartilage is a hydrated biomacromolecular fiber composite located at the ends of long bones that enables proper joint lubrication, articulation, loading, and energy dissipation. Degradation of extracellular matrix molecular components and changes in their nanoscale structure greatly influence the macroscale behavior of the tissue and result in dysfunction with age, injury, and diseases such as osteoarthritis. Here, the application of the field of nanomechanics to cartilage is reviewed. Nanomechanics involves the measurement and prediction of nanoscale forces and displacements, intra- and intermolecular interactions, spatially varying mechanical properties, and other mechanical phenomena existing at small length scales. Experimental nanomechanics and theoretical nanomechanics have been applied to cartilage at varying levels of material complexity, e.g., nanoscale properties of intact tissue, the matrix associated with single cells, biomimetic molecular assemblies, and individual extracellular matrix biomolecules (such as aggrecan, collagen, and hyaluronan). These studies have contributed to establishing a fundamental mechanism-based understanding of native and engineered cartilage tissue function, quality, and pathology.

  5. Extracellular Matrix Molecules: Potential Targets in Pharmacotherapy

    PubMed Central

    Järveläinen, Hannu; Sainio, Annele; Koulu, Markku; Wight, Thomas N.; Penttinen, Risto

    2009-01-01

    The extracellular matrix (ECM) consists of numerous macromolecules classified traditionally into collagens, elastin, and microfibrillar proteins, proteoglycans including hyaluronan, and noncollagenous glycoproteins. In addition to being necessary structural components, ECM molecules exhibit important functional roles in the control of key cellular events such as adhesion, migration, proliferation, differentiation, and survival. Any structural inherited or acquired defect and/or metabolic disturbance in the ECM may cause cellular and tissue alterations that can lead to the development or progression of disease. Consequently, ECM molecules are important targets for pharmacotherapy. Specific agents that prevent theexcess accumulation of ECM molecules in the vascular system, liver, kidney, skin, and lung; alternatively, agents that inhibit the degradation of the ECM in degenerative diseases such as osteoarthritis would be clinically beneficial. Unfortunately, until recently, the ECM in drug discovery has been largely ignored. However, several of today's drugs that act on various primary targets affect the ECM as a byproduct of the drugs' actions, and this activity may in part be beneficial to the drugs' disease-modifying properties. In the future, agents and compounds targeting directly the ECM will significantly advance the treatment of various human diseases, even those for which efficient therapies are not yet available. PMID:19549927

  6. Extracellular matrix mechanics in lung parenchymal diseases

    PubMed Central

    Suki, Béla; Bates, Jason H.T.

    2008-01-01

    In this review, we examine how the extracellular matrix (ECM) of the lung contributes to the overall mechanical properties of the parenchyma, and how these properties change in disease. The connective tissues of the lung are composed of cells and ECM, which includes a variety of biological macromolecules and water. The macromolecules that are most important in determining the mechanical properties of the ECM are collagen, elastin, and proteoglycans. We first discuss the various components of the ECM and how their architectural organization gives rise to the mechanical properties of the parenchyma. Next, we examine how mechanical forces can affect the physiological functioning of the lung parenchyma. Collagen plays an especially important role in determining the homeostasis and cellular responses to injury because it is the most important load-bearing component of the parenchyma. We then demonstrate how the concept of percolation can be used to link microscopic pathologic alterations in the parenchyma to clinically measurable lung function during the progression of emphysema and fibrosis. Finally, we speculate about the possibility of using targeted tissue engineering to optimize treatment of these two major lung diseases. PMID:18485836

  7. Instructive Roles of Extracellular Matrix on Autophagy

    PubMed Central

    Neill, Thomas; Schaefer, Liliana; Iozzo, Renato V.

    2015-01-01

    Autophagy plays an essential role in maintaining an intricate balance between nutrient demands and energetic requirements during normal homeostasis. Autophagy recycles metabolic substrates from nonspecific bulk degradation of proteins and excess or damaged organelles. Recent work posits an active and dynamic signaling role for extracellular matrix-evoked autophagic regulation, that is, allosteric and independent of prevailing nutrient conditions. Several candidates, representing a diverse repertoire of matrix constituents (decorin, collagen VI, laminin α2, endostatin, endorepellin, and kringle V), can modulate autophagic signaling pathways. Importantly, a novel principle indicates that matrix constituents can differentially modulate autophagic induction and repression via interaction with specific receptors. Most of the matrix-derived factors described here appear to control autophagy in a canonical manner but independent of nutrient deprivation. Because the molecular composition and structure of the extracellular matrix are dynamically remodeled during various physiological and pathological conditions, we propose that matrix-regulated autophagy is key for maintaining proper tissue homeostasis and disease prevention, such as cancer progression and muscular dystrophies. PMID:24976620

  8. Lung extracellular matrix and redox regulation.

    PubMed

    Watson, Walter H; Ritzenthaler, Jeffrey D; Roman, Jesse

    2016-08-01

    Pulmonary fibrosis affects millions worldwide and, even though there has been a significant investment in understanding the processes involved in wound healing and maladaptive repair, a complete understanding of the mechanisms responsible for lung fibrogenesis eludes us, and interventions capable of reversing or halting disease progression are not available. Pulmonary fibrosis is characterized by the excessive expression and uncontrolled deposition of extracellular matrix (ECM) proteins resulting in erosion of the tissue structure. Initially considered an 'end-stage' process elicited after injury, these events are now considered pathogenic and are believed to contribute to the course of the disease. By interacting with integrins capable of signal transduction and by influencing tissue mechanics, ECM proteins modulate processes ranging from cell adhesion and migration to differentiation and growth factor expression. In doing so, ECM proteins help orchestrate complex developmental processes and maintain tissue homeostasis. However, poorly controlled deposition of ECM proteins promotes inflammation, fibroproliferation, and aberrant differentiation of cells, and has been implicated in the pathogenesis of pulmonary fibrosis, atherosclerosis and cancer. Considering their vital functions, ECM proteins are the target of investigation, and oxidation-reduction (redox) reactions have emerged as important regulators of the ECM. Oxidative stress invariably accompanies lung disease and promotes ECM expression directly or through the overproduction of pro-fibrotic growth factors, while affecting integrin binding and activation. In vitro and in vivo investigations point to redox reactions as targets for intervention in pulmonary fibrosis and related disorders, but studies in humans have been disappointing probably due to the narrow impact of the interventions tested, and our poor understanding of the factors that regulate these complex reactions. This review is not meant to

  9. Lung extracellular matrix and redox regulation

    PubMed Central

    Watson, Walter H.; Ritzenthaler, Jeffrey D.; Roman, Jesse

    2016-01-01

    Pulmonary fibrosis affects millions worldwide and, even though there has been a significant investment in understanding the processes involved in wound healing and maladaptive repair, a complete understanding of the mechanisms responsible for lung fibrogenesis eludes us, and interventions capable of reversing or halting disease progression are not available. Pulmonary fibrosis is characterized by the excessive expression and uncontrolled deposition of extracellular matrix (ECM) proteins resulting in erosion of the tissue structure. Initially considered an ‘end-stage’ process elicited after injury, these events are now considered pathogenic and are believed to contribute to the course of the disease. By interacting with integrins capable of signal transduction and by influencing tissue mechanics, ECM proteins modulate processes ranging from cell adhesion and migration to differentiation and growth factor expression. In doing so, ECM proteins help orchestrate complex developmental processes and maintain tissue homeostasis. However, poorly controlled deposition of ECM proteins promotes inflammation, fibroproliferation, and aberrant differentiation of cells, and has been implicated in the pathogenesis of pulmonary fibrosis, atherosclerosis and cancer. Considering their vital functions, ECM proteins are the target of investigation, and oxidation–reduction (redox) reactions have emerged as important regulators of the ECM. Oxidative stress invariably accompanies lung disease and promotes ECM expression directly or through the overproduction of pro-fibrotic growth factors, while affecting integrin binding and activation. In vitro and in vivo investigations point to redox reactions as targets for intervention in pulmonary fibrosis and related disorders, but studies in humans have been disappointing probably due to the narrow impact of the interventions tested, and our poor understanding of the factors that regulate these complex reactions. This review is not meant to

  10. Airway and Extracellular Matrix Mechanics in COPD

    PubMed Central

    Bidan, Cécile M.; Veldsink, Annemiek C.; Meurs, Herman; Gosens, Reinoud

    2015-01-01

    Chronic obstructive pulmonary disease (COPD) is one of the most common lung diseases worldwide, and is characterized by airflow obstruction that is not fully reversible with treatment. Even though airflow obstruction is caused by airway smooth muscle contraction, the extent of airway narrowing depends on a range of other structural and functional determinants that impact on active and passive tissue mechanics. Cells and extracellular matrix in the airway and parenchymal compartments respond both passively and actively to the mechanical stimulation induced by smooth muscle contraction. In this review, we summarize the factors that regulate airway narrowing and provide insight into the relative contributions of different constituents of the extracellular matrix and their biomechanical impact on airway obstruction. We then review the changes in extracellular matrix composition in the airway and parenchymal compartments at different stages of COPD, and finally discuss how these changes impact airway narrowing and the development of airway hyperresponsiveness. Finally, we position these data in the context of therapeutic research focused on defective tissue repair. As a conclusion, we propose that future works should primarily target mild or early COPD, prior to the widespread structural changes in the alveolar compartment that are more characteristic of severe COPD. PMID:26696894

  11. Regulation of Osteoblast Survival by the Extracellular Matrix and Gravity

    NASA Technical Reports Server (NTRS)

    Globus. Ruth K.; Almeida, Eduardo A. C.; Searby, Nancy D.; Bowley, Susan M. (Technical Monitor)

    2000-01-01

    Spaceflight adversely affects the skeleton, posing a substantial risk to astronaut's health during long duration missions. The reduced bone mass observed in growing animals following spaceflight is due at least in part to inadequate bone formation by osteoblasts. Thus, it is of central importance to identify basic cellular mechanisms underlying normal bone formation. The fundamental ideas underlying our research are that interactions between extracellular matrix proteins, integrin adhesion receptors, cytoplasmic signaling and cytoskeletal proteins are key ingredients for the proper functioning of osteoblasts, and that gravity impacts these interactions. As an in vitro model system we used primary fetal rat calvarial cells which faithfully recapitulate osteoblast differentiation characteristically observed in vivo. We showed that specific integrin receptors ((alpha)3(beta)1), ((alpha)5(beta)1), ((alpha)8(betal)1) and extracellular matrix proteins (fibronectin, laminin) were needed for the differentiation of immature osteoblasts. In the course of maturation, cultured osteoblasts switched from depending on fibronectin and laminin for differentiation to depending on these proteins for their very survival. Furthermore, we found that manipulating the gravity vector using ground-based models resulted in activation of key intracellular survival signals generated by integrin/extracellular matrix interactions. We are currently testing the in vivo relevance of some of these observations using targeted transgenic technology. In conclusion, mechanical factors including gravity may participate in regulating survival via cellular interactions with the extracellular matrix. This leads us to speculate that microgravity adversely affects the survival of osteoblasts and contributes to spaceflight-induced osteoporosis.

  12. Extracellular matrix of the developing ovarian follicle.

    PubMed

    Irving-Rodgers, Helen F; Rodgers, Raymond J

    2006-09-01

    There are many different types of extracellular matrices in the different follicle compartments. These have different roles in follicle development and atresia, and they change in composition during these processes. This review focuses on basal lamina matrix in particular, and considers follicular fluid, the newly identified focimatrix, and thecal matrices. When follicles commence growing, the follicular basal lamina changes in its composition from containing all six alpha chains of type IV collagen to only alpha1 and alpha2. Perlecan and nidogen-1 and -2 subsequently become components of the follicular basal lamina, and there is an increase in the amount of laminin chains alpha1, beta2, and gamma1, in the bovine at least. Late in follicular development and on atresia some follicles contain laminin alpha2. On atresia the follicular basal lamina is not degraded, as occurs in ovulation, but can be breached by cells from the thecal layer when it is not aligned by granulosa cells. A novel type of basal lamina-like matrix, called focimatrix (abbreviated from focal intraepithelial matrix), develops between the cells of the membrana granulosa as aggregates of basal lamina material. It does not envelop cells and so cannot perform functions of basal lamina as currently understood. It is hypothesized that focimatrix assists or initiates depolarization of the membrana granulosa necessary for the transformation into luteal cells. The largest osmotically active molecules in follicular fluid are hyaluronan and chondroitin sulfate proteoglycans, including versican and inter-alpha trypsin inhibitor. It has been suggested that these might be responsible for the formation of follicular fluid by creating an osmotic gradient across the follicular wall. The formation, development, and then either ovulation or regression of follicles requires considerable tissue remodeling, cellular replication, and specialization. The expectation of researchers is that extracellular matrix will be

  13. Regulation of Corneal Stroma Extracellular Matrix Assembly

    PubMed Central

    Chen, Shoujun; Mienaltowski, Michael J.; Birk, David E.

    2014-01-01

    The transparent cornea is the major refractive element of the eye. A finely controlled assembly of the stromal extracellular matrix is critical to corneal function, as well as in establishing the appropriate mechanical stability required to maintain corneal shape and curvature. In the stroma, homogeneous, small diameter collagen fibrils, regularly packed with a highly ordered hierarchical organization, are essential for function. This review focuses on corneal stroma assembly and the regulation of collagen fibrillogenesis. Corneal collagen fibrillogenesis involves multiple molecules interacting in sequential steps, as well as interactions between keratocytes and stroma matrix components. The stroma has the highest collagen V:I ratio in the body. Collagen V regulates the nucleation of protofibril assembly, thus controlling the number of fibrils and assembly of smaller diameter fibrils in the stroma. The corneal stroma is also enriched in small leucine-rich proteoglycans (SLRPs) that cooperate in a temporal and spatial manner to regulate linear and lateral collagen fibril growth. In addition, the fibril-associated collagens (FACITs) such as collagen XII and collagen XIV have roles in the regulation of fibril packing and inter-lamellar interactions. A communicating keratocyte network contributes to the overall and long-range regulation of stromal extracellular matrix assembly, by creating micro-domains where the sequential steps in stromal matrix assembly are controlled. Keratocytes control the synthesis of extracellular matrix components, which interact with the keratocytes dynamically to coordinate the regulatory steps into a cohesive process. Mutations or deficiencies in stromal regulatory molecules result in altered interactions and deficiencies in both transparency and refraction, leading to corneal stroma pathobiology such as stromal dystrophies, cornea plana and keratoconus. PMID:25819456

  14. Achondrogenesis type II, abnormalities of extracellular matrix.

    PubMed

    Horton, W A; Machado, M A; Chou, J W; Campbell, D

    1987-09-01

    Immune and lectin histochemical and microchemical methods were employed to study growth cartilage from seven cases of achondrogenesis type II (Langer-Saldino). The normal architecture of the epiphyseal and growth plate cartilage was replaced by a morphologically heterogeneous tissue. Some areas were comprised of vascular canals surrounded by extensive fibrous tissue and enlarged cells that had the appearance and histochemical characteristics of hypertrophic chondrocytes. Other areas contained a mixture of cells ranging from small to the enlarged chondrocytes. The extracellular matrix in the latter areas was more abundant and had characteristics of both precartilage mesenchymal matrix and typical cartilage matrix; it contained types I and II collagen, cartilage proteoglycan, fibronectin, and peanut agglutinin binding glycoconjugate(s). Peptide mapping of cyanogen bromide cartilage collagen peptides revealed the presence of types I and II collagen. These observations could be explained by a defect in the biosynthesis of type II collagen or in chondrocyte differentiation. PMID:3309860

  15. Nanomechanics of the Cartilage Extracellular Matrix

    PubMed Central

    Han, Lin; Grodzinsky, Alan J.; Ortiz, Christine

    2012-01-01

    Cartilage is a hydrated biomacromolecular fiber composite located at the ends of long bones that enables proper joint lubrication, articulation, loading, and energy dissipation. Degradation of extracellular matrix molecular components and changes in their nanoscale structure greatly influence the macroscale behavior of the tissue and result in dysfunction with age, injury, and diseases such as osteoarthritis. Here, the application of the field of nanomechanics to cartilage is reviewed. Nanomechanics involves the measurement and prediction of nanoscale forces and displacements, intra- and intermolecular interactions, spatially varying mechanical properties, and other mechanical phenomena existing at small length scales. Experimental nanomechanics and theoretical nanomechanics have been applied to cartilage at varying levels of material complexity, e.g., nanoscale properties of intact tissue, the matrix associated with single cells, biomimetic molecular assemblies, and individual extracellular matrix biomolecules (such as aggrecan, collagen, and hyaluronan). These studies have contributed to establishing a fundamental mechanism-based understanding of native and engineered cartilage tissue function, quality, and pathology. PMID:22792042

  16. Bidirectional extracellular matrix signaling during tissue morphogenesis

    PubMed Central

    Gjorevski, Nikolce; Nelson, Celeste M.

    2009-01-01

    Normal tissue development and function are regulated by the interplay between cells and their surrounding extracellular matrix (ECM). The ECM provides biochemical and mechanical contextual information that is conveyed from the cell membrane through the cytoskeleton to the nucleus to direct cell phenotype. Cells, in turn, remodel the ECM and thereby sculpt their local microenvironment. Here we review the mechanisms by which cells interact with, respond to, and influence the ECM, with particular emphasis placed on the role of this bidirectional communication during tissue morphogenesis. We also discuss the implications for successful engineering of functional tissues ex vivo. PMID:19896886

  17. Extracellular Matrix Roles During Cardiac Repair

    PubMed Central

    Jourdan-LeSaux, Claude; Zhang, Jianhua; Lindsey, Merry L.

    2010-01-01

    The cardiac extracellular matrix (ECM) provides a platform for cells to maintain structure and function, which in turn maintains tissue function. In response to injury, the ECM undergoes remodeling that involves synthesis, incorporation, and degradation of matrix proteins, with the net outcome determined by the balance of these processes. The major goals of this review are a) to serve as an initial resource for students and investigators new to the cardiac ECM remodeling field, and b) to highlight a few of the key exciting avenues and methodologies that have recently been explored. While we focus on cardiac injury and responses of the left ventricle (LV), the mechanisms reviewed here have pathways in common with other wound healing models. PMID:20670633

  18. Tendon Extracellular Matrix Alterations in Ullrich Congenital Muscular Dystrophy.

    PubMed

    Sardone, Francesca; Traina, Francesco; Bondi, Alice; Merlini, Luciano; Santi, Spartaco; Maraldi, Nadir Mario; Faldini, Cesare; Sabatelli, Patrizia

    2016-01-01

    Collagen VI (COLVI) is a non-fibrillar collagen expressed in skeletal muscle and most connective tissues. Mutations in COLVI genes cause two major clinical forms, Bethlem myopathy and Ullrich congenital muscular dystrophy (UCMD). In addition to congenital muscle weakness, patients affected by COLVI myopathies show axial and proximal joint contractures and distal joint hypermobility, which suggest the involvement of the tendon function. We examined a peroneal tendon biopsy and tenocyte culture of a 15-year-old patient affected by UCMD with compound heterozygous COL6A2 mutations. In patient's tendon biopsy, we found striking morphological alterations of tendon fibrils, consisting in irregular profiles and reduced mean diameter. The organization of the pericellular matrix of tenocytes, the primary site of collagen fibril assembly, was severely affected, as determined by immunoelectron microscopy, which showed an abnormal accumulation of COLVI and altered distribution of collagen I (COLI) and fibronectin (FBN). In patient's tenocyte culture, COLVI web formation and cell surface association were severely impaired; large aggregates of COLVI, which matched with COLI labeling, were frequently detected in the extracellular matrix. In addition, metalloproteinase MMP-2, an extracellular matrix-regulating enzyme, was increased in the conditioned medium of patient's tenocytes, as determined by gelatin zymography and western blot. Altogether, these data indicate that COLVI deficiency may influence the organization of UCMD tendon matrix, resulting in dysfunctional fibrillogenesis. The alterations of tendon matrix may contribute to the complex pathogenesis of COLVI related myopathies. PMID:27375477

  19. Tendon Extracellular Matrix Alterations in Ullrich Congenital Muscular Dystrophy

    PubMed Central

    Sardone, Francesca; Traina, Francesco; Bondi, Alice; Merlini, Luciano; Santi, Spartaco; Maraldi, Nadir Mario; Faldini, Cesare; Sabatelli, Patrizia

    2016-01-01

    Collagen VI (COLVI) is a non-fibrillar collagen expressed in skeletal muscle and most connective tissues. Mutations in COLVI genes cause two major clinical forms, Bethlem myopathy and Ullrich congenital muscular dystrophy (UCMD). In addition to congenital muscle weakness, patients affected by COLVI myopathies show axial and proximal joint contractures and distal joint hypermobility, which suggest the involvement of the tendon function. We examined a peroneal tendon biopsy and tenocyte culture of a 15-year-old patient affected by UCMD with compound heterozygous COL6A2 mutations. In patient’s tendon biopsy, we found striking morphological alterations of tendon fibrils, consisting in irregular profiles and reduced mean diameter. The organization of the pericellular matrix of tenocytes, the primary site of collagen fibril assembly, was severely affected, as determined by immunoelectron microscopy, which showed an abnormal accumulation of COLVI and altered distribution of collagen I (COLI) and fibronectin (FBN). In patient’s tenocyte culture, COLVI web formation and cell surface association were severely impaired; large aggregates of COLVI, which matched with COLI labeling, were frequently detected in the extracellular matrix. In addition, metalloproteinase MMP-2, an extracellular matrix-regulating enzyme, was increased in the conditioned medium of patient’s tenocytes, as determined by gelatin zymography and western blot. Altogether, these data indicate that COLVI deficiency may influence the organization of UCMD tendon matrix, resulting in dysfunctional fibrillogenesis. The alterations of tendon matrix may contribute to the complex pathogenesis of COLVI related myopathies. PMID:27375477

  20. [Hematopoietic microenvironment: cellular and extracellular matrix elements].

    PubMed

    Minguell, J J; Fernández, M; Tetas, M; Martínez, J; Bruzzone, M; Rodríguez, J P

    1988-06-01

    In bone marrow, cellular stroma together with extracellular matrix (EM) provide an adequate microenvironment for the proliferation and differentiation of hemopoietic progenitor cells. In this article we describe studies on the cell characteristics of a main stromal phenotype, a fibroblast-like cell and its ability to produce in vitro EM components. Comparative studies were performed in fibroblast cultures derived from normal and acute lymphoblastic leukemic (ALL) bone marrow. The grow characteristics of fibroblasts from ALL marrow as well as its capacity to synthetize collagen, fibronectin and GAGs are impaired when compared to fibroblast from normal marrow. Thus, in ALL the impaired production of EM biomolecules by a transient damaged population of stromal cells, may contribute to the development of a defective microenvironment for hemopoiesis. PMID:3154858

  1. Extracellular matrix motion and early morphogenesis.

    PubMed

    Loganathan, Rajprasad; Rongish, Brenda J; Smith, Christopher M; Filla, Michael B; Czirok, Andras; Bénazéraf, Bertrand; Little, Charles D

    2016-06-15

    For over a century, embryologists who studied cellular motion in early amniotes generally assumed that morphogenetic movement reflected migration relative to a static extracellular matrix (ECM) scaffold. However, as we discuss in this Review, recent investigations reveal that the ECM is also moving during morphogenesis. Time-lapse studies show how convective tissue displacement patterns, as visualized by ECM markers, contribute to morphogenesis and organogenesis. Computational image analysis distinguishes between cell-autonomous (active) displacements and convection caused by large-scale (composite) tissue movements. Modern quantification of large-scale 'total' cellular motion and the accompanying ECM motion in the embryo demonstrates that a dynamic ECM is required for generation of the emergent motion patterns that drive amniote morphogenesis. PMID:27302396

  2. The Extracellular Matrix and Insulin Resistance

    PubMed Central

    Williams, Ashley S.; Kang, Li; Wasserman, David H.

    2015-01-01

    The extracellular matrix (ECM) is a highly dynamic compartment that undergoes remodeling as a result of injury and repair. Over the past decade, mounting evidence in humans and rodents suggest that ECM remodeling is associated with diet-induced insulin resistance in several metabolic tissues. Additionally, integrin receptors for the ECM have also been implicated in the regulation of insulin action. This review will address what is currently known about the ECM, integrins and insulin action in the muscle, liver and adipose tissue. Understanding how ECM remodeling and integrin signaling regulates insulin action may aid in the development of new therapeutic targets for the treatment of insulin resistance and type 2 diabetes. PMID:26059707

  3. Extracellular Matrix Revisited: Roles in Tissue Engineering

    PubMed Central

    2016-01-01

    The extracellular matrix (ECM) is a heterogeneous, connective network composed of fibrous glycoproteins that coordinate in vivo to provide the physical scaffolding, mechanical stability, and biochemical cues necessary for tissue morphogenesis and homeostasis. This review highlights some of the recently raised aspects of the roles of the ECM as related to the fields of biophysics and biomedical engineering. Fundamental aspects of focus include the role of the ECM as a basic cellular structure, for novel spontaneous network formation, as an ideal scaffold in tissue engineering, and its essential contribution to cell sheet technology. As these technologies move from the laboratory to clinical practice, they are bound to shape the vast field of tissue engineering for medical transplantations. PMID:27230457

  4. Extracellular matrix fluctuations during early embryogenesis

    PubMed Central

    Szabó, A; Rupp, P A; Rongish, B J; Little, C D; Czirók, A

    2011-01-01

    Extracellular matrix (ECM) movements and rearrangements were studied in avian embryos during early stages of development. We show that the ECM moves as a composite material, whereby distinct molecular components as well as spatially separated layers exhibit similar displacements. Using scanning wide field and confocal microscopy we show that the velocity field of ECM displacement is smooth in space and that ECM movements are correlated even at locations separated by several hundred micrometers. Velocity vectors, however, strongly fluctuate in time. The autocorrelation time of the velocity fluctuations is less than a minute. Suppression of the fluctuations yields a persistent movement pattern that is shared among embryos at equivalent stages of development. The high resolution of the velocity fields allows a detailed spatio-temporal characterization of important morphogenetic processes, especially tissue dynamics surrounding the embryonic organizer (Hensen’s node). PMID:21750366

  5. Extracellular matrix and pathogenic mechanisms in osteoarthritis.

    PubMed

    Hardingham, Tim

    2008-01-01

    Osteoarthritis (OA) is a heterogeneous condition of joint degeneration characterized by structural changes in extracellular matrices such as subchondral bone and cartilage. Research has identified many diverse ways of initiating OA, varying from mechanical disruption to gene mutations in structural proteins. A frequent end point is cartilage loss, which can occur irrespective of the initiating mechanism. Of the mechanisms responsible for cartilage matrix damage, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-5 was identified as of key importance in knockout mice, but work with human cartilage has suggested that ADAMTS-4 was also involved. A transgenic mouse expressing aggrecan lacking a key aggrecanase site clearly showed that loss of aggrecan from cartilage was an important step in both inflammatory and trauma-induced joint degeneration. In OA, cartilage chondrocytes show changes in gene expression, and it remains to be resolved if this reflects adaptive responses to changes in biological, physical, and mechanical signaling rather than any form of differentiation. PMID:18457609

  6. Defining the extracellular matrix using proteomics

    PubMed Central

    Byron, Adam; Humphries, Jonathan D; Humphries, Martin J

    2013-01-01

    The cell microenvironment has a profound influence on the behaviour, growth and survival of cells. The extracellular matrix (ECM) provides not only mechanical and structural support to cells and tissues but also binds soluble ligands and transmembrane receptors to provide spatial coordination of signalling processes. The ability of cells to sense the chemical, mechanical and topographical features of the ECM enables them to integrate complex, multiparametric information into a coherent response to the surrounding microenvironment. Consequently, dysregulation or mutation of ECM components results in a broad range of pathological conditions. Characterization of the composition of ECM derived from various cells has begun to reveal insights into ECM structure and function, and mechanisms of disease. Proteomic methodologies permit the global analysis of subcellular systems, but extracellular and transmembrane proteins present analytical difficulties to proteomic strategies owing to the particular biochemical properties of these molecules. Here, we review advances in proteomic approaches that have been applied to furthering our understanding of the ECM microenvironment. We survey recent studies that have addressed challenges in the analysis of ECM and discuss major outcomes in the context of health and disease. In addition, we summarize efforts to progress towards a systems-level understanding of ECM biology. PMID:23419153

  7. Extracellular Matrix Dynamics and Fetal Membrane Rupture

    PubMed Central

    Strauss,, Jerome F.

    2013-01-01

    The extracellular matrix (ECM) plays an important role in determining cell and organ function: (1) it is an organizing substrate that provides tissue tensile strength; (2) it anchors cells and influences cell morphology and function via interaction with cell surface receptors; and (3) it is a reservoir for growth factors. Alterations in the content and the composition of the ECM determine its physical and biological properties, including strength and susceptibility to degradation. The ECM components themselves also harbor cryptic matrikines, which when exposed by conformational change or proteolysis have potent effects on cell function, including stimulating the production of cytokines and matrix metalloproteinases (MMPs). Collectively, these properties of the ECM reflect a dynamic tissue component that influences both tissue form and function. This review illustrates how defects in ECM synthesis and metabolism and the physiological process of ECM turnover contribute to changes in the fetal membranes that precede normal parturition and contribute to the pathological events leading to preterm premature rupture of membranes (PPROM). PMID:22267536

  8. Extracellular matrix dynamics and fetal membrane rupture.

    PubMed

    Strauss, Jerome F

    2013-02-01

    The extracellular matrix (ECM) plays an important role in determining cell and organ function: (1) it is an organizing substrate that provides tissue tensile strength; (2) it anchors cells and influences cell morphology and function via interaction with cell surface receptors; and (3) it is a reservoir for growth factors. Alterations in the content and the composition of the ECM determine its physical and biological properties, including strength and susceptibility to degradation. The ECM components themselves also harbor cryptic matrikines, which when exposed by conformational change or proteolysis have potent effects on cell function, including stimulating the production of cytokines and matrix metalloproteinases (MMPs). Collectively, these properties of the ECM reflect a dynamic tissue component that influences both tissue form and function. This review illustrates how defects in ECM synthesis and metabolism and the physiological process of ECM turnover contribute to changes in the fetal membranes that precede normal parturition and contribute to the pathological events leading to preterm premature rupture of membranes (PPROM). PMID:22267536

  9. Roles of extracellular matrix in follicular development.

    PubMed

    Rodgers, R J; van Wezel, I L; Irving-Rodgers, H F; Lavranos, T C; Irvine, C M; Krupa, M

    1999-01-01

    The cellular biology and changes in the extracellular matrix of ovarian follicles during their development are reviewed. During growth of the bovine ovarian follicle the follicular basal lamina doubles 19 times in surface area. It changes in composition, having collagen IV alpha 1-26 and laminin alpha 1, beta 2 and gamma 1 at the primordial stage, and collagen IV alpha 1 and alpha 2, reduced amounts of alpha 3-alpha 5, and a higher content of laminin alpha 1, beta 2 and gamma 1 at the antral stage. In atretic antral follicles laminin alpha 2 was also detected. The follicular epithelium also changes from one layer to many layers during follicular growth. It is clear that not all granulosal cells have equal potential to divide, and we have evidence that the granulosal cells arise from a population of stem cells. This finding has important ramifications and supports the concept that different follicular growth factors can act on different subsets of granulosal cells. In antral follicles, the replication of cells occurs in the middle layers of the membrana granulosa, with older granulosal cells towards the antrum and towards the basal lamina. The basal cells in the membrana granulosa have also been observed to vary in shape between follicies. In smaller antral follicles, they were either columnar or rounded, and in follicles > 5 mm the cells were all rounded. The reasons for these changes in matrix and cell shapes are discussed in relation to follicular development. PMID:10692866

  10. Tumorigenic potential of extracellular matrix metalloproteinase inducer.

    PubMed

    Zucker, S; Hymowitz, M; Rollo, E E; Mann, R; Conner, C E; Cao, J; Foda, H D; Tompkins, D C; Toole, B P

    2001-06-01

    Extracellular matrix metalloproteinase inducer (EMMPRIN), a glycoprotein present on the cancer cell plasma membrane, enhances fibroblast synthesis of matrix metalloproteinases (MMPs). The demonstration that peritumoral fibroblasts synthesize most of the MMPs in human tumors rather than the cancer cells themselves has ignited interest in the role of EMMPRIN in tumor dissemination. In this report we have demonstrated a role for EMMPRIN in cancer progression. Human MDA-MB-436 breast cancer cells, which are tumorigenic but slow growing in vivo, were transfected with EMMPRIN cDNA and injected orthotopically into mammary tissue of female NCr nu/nu mice. Green fluorescent protein was used to visualize metastases. In three experiments, breast cancer cell clones transfected with EMMPRIN cDNA were considerably more tumorigenic and invasive than plasmid-transfected cancer cells. Increased gelatinase A and gelatinase B expression (demonstrated by in situ hybridization and gelatin substrate zymography) was demonstrated in EMMPRIN-enhanced tumors. In contrast to de novo breast cancers in humans, human tumors transplanted into mice elicited minimal stromal or inflammatory cell reactions. Based on these experimental studies and our previous demonstration that EMMPRIN is prominently displayed in human cancer tissue, we propose that EMMPRIN plays an important role in cancer progression by increasing synthesis of MMPs. PMID:11395366

  11. Interactions of Cartilage Extracellular Matrix Macromolecules.

    PubMed

    Horkay, Ferenc

    2012-12-15

    Articular cartilage is a low-friction, load-bearing tissue located at joint surfaces. The extracellular matrix (ECM) of cartilage consists of a fibrous collagen network, which is pre-stressed by the osmotic swelling pressure exerted by negatively charged proteoglycan aggregates embedded in the collagen network. The major proteoglycan is the bottlebrush shaped aggrecan, which forms complexes with linear hyaluronic acid chains. We quantify microscopic and macroscopic changes resulting from self-assembly between aggrecan and hyaluronic acid using a complementary set of physical measurements to determine structure and interactions by combining scattering techniques, including small-angle X-ray scattering, small-angle neutron scattering, and dynamic light scattering with macroscopic osmotic pressure measurements. It is demonstrated that the osmotic pressure that defines the load bearing ability of cartilage is primarily governed by the main macromolecular components (aggrecan and collagen) of the ECM. Knowledge of the interactions between the macromolecular components of cartilage ECM is essential to understand biological function and to develop successful tissue engineering strategies for cartilage repair. PMID:23997426

  12. Vascular Extracellular Matrix and Arterial Mechanics

    PubMed Central

    WAGENSEIL, JESSICA E.; MECHAM, ROBERT P.

    2009-01-01

    An important factor in the transition from an open to a closed circulatory system was a change in vessel wall structure and composition that enabled the large arteries to store and release energy during the cardiac cycle. The component of the arterial wall in vertebrates that accounts for these properties is the elastic fiber network organized by medial smooth muscle. Beginning with the onset of pulsatile blood flow in the developing aorta, smooth muscle cells in the vessel wall produce a complex extracellular matrix (ECM) that will ultimately define the mechanical properties that are critical for proper function of the adult vascular system. This review discusses the structural ECM proteins in the vertebrate aortic wall and will explore how the choice of ECM components has changed through evolution as the cardiovascular system became more advanced and pulse pressure increased. By correlating vessel mechanics with physiological blood pressure across animal species and in mice with altered vessel compliance, we show that cardiac and vascular development are physiologically coupled, and we provide evidence for a universal elastic modulus that controls the parameters of ECM deposition in vessel wall development. We also discuss mechanical models that can be used to design better tissue-engineered vessels and to test the efficacy of clinical treatments. PMID:19584318

  13. Surface Characterization of Extracellular Matrix Scaffolds

    PubMed Central

    Brown, Bryan N.; Barnes, Christopher A.; Kasick, Rena T.; Michel, Roger; Gilbert, Thomas W.; Beer-Stolz, Donna; Castner, David G.; Ratner, Buddy D.; Badylak, Stephen F.

    2009-01-01

    Extracellular matrix (ECM) scaffolds prepared from different tissue sources or using different methods have been demonstrated to have distinctive effects upon cell adhesion patterns and the ability to support and maintain differentiated phenotypes. It is unknown whether the molecular composition or the ultrastructure of the ECM plays a greater role in determining the phenotype of the cells with which it comes into contact. However, when implanted, the topology and ligand landscape of the material will determine the host molecules that bind and the type and behavior of cells that mediate the host response. Therefore, a comprehensive understanding of surface characteristics is essential in the design of scaffolds for specific clinical applications. The surface characteristics of ECM scaffolds derived from porcine urinary bladder, small intestine, and liver as well as the effects of two commonly used methods of chemical cross-linking upon UBM were investigated. Electron microscopy and time of flight secondary ion mass spectroscopy were used to examine the surface characteristics of the scaffolds. The results show that ECM scaffolds have unique morphologic and structural properties which are dependant on the organ or tissue from which the scaffold is harvested. Furthermore, the results show that the surface characteristics of an ECM scaffold are changed through chemical cross-linking. PMID:19828192

  14. Biochemistry of the extracellular matrix of Volvox.

    PubMed

    Sumper, M; Hallmann, A

    1998-01-01

    The volvocine algae range in complexity from unicellular Chlamydomonas to multicellular organisms in the genus Volvox. The transition from unicellularity to multicellularity in the Volvocales is a recent event in evolution. Thus, these organisms provide a unique opportunity for exploring the development of a complex extracellular matrix (ECM) from the cell wall of a unicellular ancestor. The ECM of Volvox is divided into four main zones: The flagellar, boundary, cellular, and deep zones. Each zone is defined by ultrastructure and by characteristic ECM glycoproteins. Volvox ECM is modified under developmental control or in response to external stimuli, like the sex-inducing pheromone or stress factors. The structures of more than 10 ECM glycoproteins from a single species of Volvox are now known in molecular detail and are compared to other algal and plant cell wall/ECM glycoproteins. Although usually classified as hydroxyproline-rich glycoproteins, the striking feature of all algal ECM glycoproteins is a modular composition. Rod-shaped hydroxyproline-rich modules are combined with hydroxyproline-free domains that meet the multiple functional requirements of a complex ECM. The algal ECM provides another example of the combinatorial advantage of shuffling modules that is so evident in the evolution of the metazoan ECMs. PMID:9496634

  15. Lung Extracellular Matrix and Fibroblast Function

    PubMed Central

    2015-01-01

    Extracellular matrix (ECM) is a tissue-specific macromolecular structure that provides physical support to tissues and is essential for normal organ function. In the lung, ECM plays an active role in shaping cell behavior both in health and disease by virtue of the contextual clues it imparts to cells. Qualities including dimensionality, molecular composition, and intrinsic stiffness all promote normal function of the lung ECM. Alterations in composition and/or modulation of stiffness of the focally injured or diseased lung ECM microenvironment plays a part in reparative processes performed by fibroblasts. Under conditions of remodeling or in disease states, inhomogeneous stiffening (or softening) of the pathologic ECM may both precede modifications in cell behavior and be a result of disease progression. The ability of ECM to stimulate further ECM production by fibroblasts and drive disease progression has potentially significant implications for mesenchymal stromal cell–based therapies; in the setting of pathologic ECM stiffness or composition, the therapeutic intent of progenitor cells may be subverted. Taken together, current data suggest that lung ECM actively contributes to health and disease; thus, mediators of cell–ECM signaling or factors that influence ECM stiffness may represent viable therapeutic targets in many lung disorders. PMID:25830832

  16. Engineering hydrogels as extracellular matrix mimics

    PubMed Central

    Geckil, Hikmet; Xu, Feng; Zhang, Xiaohui; Moon, SangJun

    2010-01-01

    Extracellular matrix (ECM) is a complex cellular environment consisting of proteins, proteoglycans, and other soluble molecules. ECM provides structural support to mammalian cells and a regulatory milieu with a variety of important cell functions, including assembling cells into various tissues and organs, regulating growth and cell–cell communication. Developing a tailored in vitro cell culture environment that mimics the intricate and organized nanoscale meshwork of native ECM is desirable. Recent studies have shown the potential of hydrogels to mimic native ECM. Such an engineered native-like ECM is more likely to provide cells with rational cues for diagnostic and therapeutic studies. The research for novel biomaterials has led to an extension of the scope and techniques used to fabricate biomimetic hydrogel scaffolds for tissue engineering and regenerative medicine applications. In this article, we detail the progress of the current state-of-the-art engineering methods to create cell-encapsulating hydrogel tissue constructs as well as their applications in in vitro models in biomedicine. PMID:20394538

  17. The extracellular matrix of plants: Molecular, cellular and developmental biology

    SciTech Connect

    1996-12-31

    A symposium entitled ``The Extracellular Matrix of Plants: Molecular, Cellular and Developmental Biology was held in Tamarron, Colorado, March 15--21, 1996. The following topics were explored in addresses by 43 speakers: structure and biochemistry of cell walls; biochemistry, molecular biology and biosynthesis of lignin; secretory pathway and synthesis of glycoproteins; biosynthesis of matrix polysaccharides, callose and cellulose; role of the extracellular matrix in plant growth and development; plant cell walls in symbiosis and pathogenesis.

  18. The extracellular matrix in hepatic regeneration.

    PubMed

    Martinez-Hernandez, A; Amenta, P S

    1995-11-01

    After partial hepatectomy, as a consequence of hepatocyte proliferation, cell clusters containing 10-14 hepatocytes are formed. These clusters are devoid of sinusoids and extracellular matrix; therefore, many hepatocytes are two to three cells removed from the vascular spaces. Four days after hepatectomy, Ito cells send delicate cell processes between the hepatocytes in the clusters. This "invasion" of the clusters coincides with the activation in Ito cells of genes encoding for several laminin chains. The penetration of Ito cells into the clusters is followed by fenestrated endothelial cells, and in this manner the normal hepatocyte vascular relationship is restored. As soon as the normal vascular structure is reestablished, the laminin genes are turned off. This chain of events is similar to the one taking place during hepatogenesis when continuous capillaries are converted into sinusoids. This similarity in hepatogenesis and regeneration suggests that the secreted laminin chains may be signals for the vascularization of the clusters by fenestrated sinusoids. During this process neither entactin nor laminin alpha chains are secreted. The vascularization of the regenerating clusters contrasts sharply to the vascularization of cirrhotic nodules. In the latter case, entactin and perhaps laminin alpha 1 chains are secreted, and the final result is the formation of basement membranes and continuous capillaries rather than fenestrated sinusoids. We suggest that entactin and specific laminin chains play a crucial role in determining the outcome of hepatic injury. Definition of the roles of entactin and laminin chains in vascularization and modulation of the endothelial phenotype will not only elucidate important aspects of regeneration, but may provide a better understanding of cirrhosis and even suggest therapeutic approaches. PMID:7589981

  19. Extracellular Matrix and Its Role in Spermatogenesis

    PubMed Central

    Siu, Michelle K.Y.; Cheng, C. Yan

    2014-01-01

    In adult mammalian testes, such as rats, Sertoli and germ cells at different stages of their development in the seminiferous epithelium are in close contact with the basement membrane, a modified form of extracellular matrix (ECM). In essence, Sertoli and germ cells in particular spermatogonia are “resting” on the basement membrane at different stages of the seminiferous epithelial cycle, relying on its structural and hormonal supports. Thus, it is not entirely unexpected that ECM plays a significant role in regulating spermatogenesis, particularly spermatogonia and Sertoli cells, and the blood-testis barrier (BTB) constituted by Sertoli cells since these cells are in physical contact with the basement membrane. Additionally, the basement membrane is also in close contact with the underlying collagen network and the myoid cell layers, which together with the lymphatic network, constitute the tunica propria. The seminiferous epithelium and the tunica propria, in turn, constitute the seminiferous tubule, which is the functional unit that produces spermatozoa via its interaction with Leydig cells in the interstitium. In short, the basement membrane and the underlying collagen network that create the acellular zone of the tunica propria may even facilitate cross-talk between the seminiferous epithelium, the myoid cells and cells in the interstitium. Recent studies in the field have illustrated the crucial role of ECM in supporting Sertoli and germ cell function in the seminiferous epithelium, including the BTB dynamics. In this chapter, we summarize some of the latest findings in the field regarding the functional role of ECM in spermatogenesis using the adult rat testis as a model. We also highlight specific areas of research that deserve attention for investigators in the field. PMID:19856163

  20. Effects of ionizing radiation on extracellular matrix

    NASA Astrophysics Data System (ADS)

    Mohamed, F.; Bradley, D. A.; Winlove, C. P.

    2007-09-01

    The extracellular matrix is a ubiquitous and important component of tissues. We investigated the effects of ionizing radiation on the physical properties of its principal macromolecular components, pericardial collagen, ligament elastin and hyaluronan, a representative glycosaminoglycan. Samples were exposed to X-rays from an electron linear accelerator in the range of 10-100 Gy to cover the range of irradiation exposure during radiotherapy. A uniaxial mechanical testing protocol was used to characterize the fibrous proteins. For pericardial tissue the major change was an increase in the elastic modulus in the toe region of the curve (⩽20% strain), from 23±18 kPa for controls to 57±22 kPa at a dose of 10 Gy ( p=0.01, α=0.05). At larger strain (⩾20% strain), the elastic modulus in the linear region decreased from 1.92±0.70 MPa for control pericardium tissue to 1.31±0.56 MPa ( p=0.01, α=0.05) for 10 Gy X-irradiated sample. Similar observations have been made previously on tendon collagen at larger strains. For elastin, the stress-strain relationship was linear up to 30% strain, but the elastic modulus decreased significantly with irradiation (controls 626±65 kPa, irradiated 474±121 kPa ( p=0.02, α=0.05), at 10 Gy X-irradiation). The results suggest that for collagen the primary effect of irradiation is generation of additional cross-links, while for elastin chain scissions are important. The viscosity of HA (at 1.25% w/v and 0.125% w/v) was measured by both cone and plate and capillary viscometry, the former providing measurement at uniform shear rate and the latter providing a more sensitive indication of changes at low viscosity. Both techniques revealed a dose-dependent reduction in viscosity (from 3400±194 cP for controls to 1500±88 cP at a shear rate of 2 s -1 and dose of 75 Gy), again suggesting depolymerization.

  1. Intermolecular interactions of thrombospondins drive their accumulation in extracellular matrix

    PubMed Central

    Kim, Dae Joong; Christofidou, Elena D.; Keene, Douglas R.; Hassan Milde, Marwah; Adams, Josephine C.

    2015-01-01

    Thrombospondins participate in many aspects of tissue organization in adult tissue homeostasis, and their dysregulation contributes to pathological processes such as fibrosis and tumor progression. The incorporation of thrombospondins into extracellular matrix (ECM) as discrete puncta has been documented in various tissue and cell biological contexts, yet the underlying mechanisms remain poorly understood. We find that collagen fibrils are disorganized in multiple tissues of Thbs1−/− mice. In investigating how thrombospondins become retained within ECM and thereby affect ECM organization, we find that accumulation of thrombospondin-1 or thrombospondin-5 puncta within cell-derived ECM is controlled by a novel, conserved, surface-exposed site on the thrombospondin L-type lectin domain. This site acts to recruit thrombospondin molecules into ECM by intermolecular interactions in trans. This mechanism is fibronectin independent, can take place extracellularly, and is demonstrated to be direct in vitro. The trans intermolecular interactions can also be heterotypic—for example, between thrombospondin-1 and thrombospondin-5. These data identify a novel concept of concentration-dependent, intermolecular “matrix trapping” as a conserved mechanism that controls the accumulation and thereby the functionality of thrombospondins in ECM. PMID:25995382

  2. Oxidation and modification of extracellular matrix and its role in disease.

    PubMed

    Chuang, C Y; Degendorfer, G; Davies, M J

    2014-09-01

    There is accumulating evidence that damage to extracellular materials and particularly the extracellular matrix, can play a major role in multiple human pathologies. In contrast to cells, the extracellular compartment of most biological tissues is relatively poorly equipped to prevent or repair damage caused by oxidation due to lower levels of antioxidant defenses (low molecular mass and enzymatic) and repair systems (both catabolic and enzymatic). The extracellular compartment is therefore likely to be subject to both an increased extent of damage and an overall accumulation of damage due to slow turnover and/or poor repair. The nature and consequences of damage to the extracellular matrix is poorly understood, despite evidence that changes in matrix structure influences not only structural integrity, but also cell adhesion, proliferation, migration and signaling, and cytokine and growth factor binding. In this article the nature of the extracellular matrix is briefly reviewed, together with evidence for the presence of matrix modifications in cardiovascular disease. The oxidants and mechanisms that are known to damage extracellular matrix are reviewed, together with the limited data available to date on how such changes affect structural properties and cellular behavior. PMID:24796988

  3. Remodelling the extracellular matrix in development and disease

    PubMed Central

    Bonnans, Caroline; Chou, Jonathan; Werb, Zena

    2015-01-01

    The extracellular matrix (ECM) is a highly dynamic structure that is present in all tissues and continuously undergoes controlled remodelling. This process involves quantitative and qualitative changes in the ECM, mediated by specific enzymes that are responsible for ECM degradation, such as metalloproteinases. The ECM interacts with cells to regulate diverse functions, including proliferation, migration and differentiation. ECM remodelling is crucial for regulating the morphogenesis of the intestine and lungs, as well as of the mammary and submandibular glands. Dysregulation of ECM composition, structure, stiffness and abundance contributes to several pathological conditions, such as fibrosis and invasive cancer. A better understanding of how the ECM regulates organ structure and function and of how ECM remodelling affects disease progression will contribute to the development of new therapeutics. PMID:25415508

  4. Extracellular matrix production in vitro in cartilage tissue engineering

    PubMed Central

    2014-01-01

    Cartilage tissue engineering is arising as a technique for the repair of cartilage lesions in clinical applications. However, fibrocartilage formation weakened the mechanical functions of the articular, which compromises the clinical outcomes. Due to the low proliferation ability, dedifferentiation property and low production of cartilage-specific extracellular matrix (ECM) of the chondrocytes, the cartilage synthesis in vitro has been one of the major limitations for obtaining high-quality engineered cartilage constructs. This review discusses cells, biomaterial scaffolds and stimulating factors that can facilitate the cartilage-specific ECM production and accumulation in the in vitro culture system. Special emphasis has been put on the factors that affect the production of ECM macromolecules such as collagen type II and proteoglycans in the review, aiming at providing new strategies to improve the quality of tissue-engineered cartilage. PMID:24708713

  5. Extracellular Matrix Modulation: Optimizing Skin Care and Rejuvenation Procedures.

    PubMed

    Widgerow, Alan D; Fabi, Sabrina G; Palestine, Roberta F; Rivkin, Alexander; Ortiz, Arisa; Bucay, Vivian W; Chiu, Annie; Naga, Lina; Emer, Jason; Chasan, Paul E

    2016-04-01

    Normal aging and photoaging of the skin are chronic processes that progress gradually. The extracellular matrix (ECM), constituting over 70% of the skin, is the central hub for repair and regeneration of the skin. As such, the ECM is the area where changes related to photodamage are most evident. Degradation of the ECM with fragmentation of proteins significantly affects cross talk and signaling between cells, the matrix, and its constituents. The accumulation of collagen fragments, amorphous elastin agglutinations, and abnormal cross-linkages between the collagen fragments impedes the ECM from its normal repair and regenerative capacity, which manifests as wrinkled, non-elastic skin. Similar to how the chronic wound healing process requires wound bed preparation before therapeutic intervention, treatment of chronic aging of the skin would likely benefit from a "skin bed preparation" to optimize the outcome of rejuvenation procedures and skin maintenance programs. This involves introducing agents that can combat stress-induced oxidation, proteasome dysfunction, and non-enzymatic cross linkages involved in glycation end products, to collectively modulate this damaged ECM, and upregulate neocollagenesis and elastin production. Agents of particular interest are matrikines, peptides originating from the fragmentation of matrix proteins that exhibit a wide range of biological activities. Peptides of this type (tripeptide and hexapeptide) are incorporated in ALASTIN™ Skin Nectar with TriHex™ technology (ALASTIN Skincare, Inc., Carlsbad, CA), which is designed to target ECM modulation with a goal of optimizing results following invasive and non-invasive dermal rejuvenating procedures. PMID:27050707

  6. The extracellular matrix modulates the hallmarks of cancer.

    PubMed

    Pickup, Michael W; Mouw, Janna K; Weaver, Valerie M

    2014-12-01

    The extracellular matrix regulates tissue development and homeostasis, and its dysregulation contributes to neoplastic progression. The extracellular matrix serves not only as the scaffold upon which tissues are organized but provides critical biochemical and biomechanical cues that direct cell growth, survival, migration and differentiation and modulate vascular development and immune function. Thus, while genetic modifications in tumor cells undoubtedly initiate and drive malignancy, cancer progresses within a dynamically evolving extracellular matrix that modulates virtually every behavioral facet of the tumor cells and cancer-associated stromal cells. Hanahan and Weinberg defined the hallmarks of cancer to encompass key biological capabilities that are acquired and essential for the development, growth and dissemination of all human cancers. These capabilities include sustained proliferation, evasion of growth suppression, death resistance, replicative immortality, induced angiogenesis, initiation of invasion, dysregulation of cellular energetics, avoidance of immune destruction and chronic inflammation. Here, we argue that biophysical and biochemical cues from the tumor-associated extracellular matrix influence each of these cancer hallmarks and are therefore critical for malignancy. We suggest that the success of cancer prevention and therapy programs requires an intimate understanding of the reciprocal feedback between the evolving extracellular matrix, the tumor cells and its cancer-associated cellular stroma. PMID:25381661

  7. The extracellular matrix modulates the hallmarks of cancer

    PubMed Central

    Pickup, Michael W; Mouw, Janna K; Weaver, Valerie M

    2014-01-01

    The extracellular matrix regulates tissue development and homeostasis, and its dysregulation contributes to neoplastic progression. The extracellular matrix serves not only as the scaffold upon which tissues are organized but provides critical biochemical and biomechanical cues that direct cell growth, survival, migration and differentiation and modulate vascular development and immune function. Thus, while genetic modifications in tumor cells undoubtedly initiate and drive malignancy, cancer progresses within a dynamically evolving extracellular matrix that modulates virtually every behavioral facet of the tumor cells and cancer-associated stromal cells. Hanahan and Weinberg defined the hallmarks of cancer to encompass key biological capabilities that are acquired and essential for the development, growth and dissemination of all human cancers. These capabilities include sustained proliferation, evasion of growth suppression, death resistance, replicative immortality, induced angiogenesis, initiation of invasion, dysregulation of cellular energetics, avoidance of immune destruction and chronic inflammation. Here, we argue that biophysical and biochemical cues from the tumor-associated extracellular matrix influence each of these cancer hallmarks and are therefore critical for malignancy. We suggest that the success of cancer prevention and therapy programs requires an intimate understanding of the reciprocal feedback between the evolving extracellular matrix, the tumor cells and its cancer-associated cellular stroma. PMID:25381661

  8. Monitoring of Extracellular Matrix Formation using Nanosecond Pulsed Laser

    NASA Astrophysics Data System (ADS)

    Ishihara, Miya; Sato, Masato; Mitani, Genya; Nagai, Toshihiro; Kutsuna, Toshiharu; Mochida, Joji; Kikuchi, Makoto

    There is a new demand in the field of tissue engineering for evaluation technology of extracellular matrix because the extracellular matrix plays an important role in the function of skeletal tissue such as articular cartilage. We previously proposed a noninvasive method of viscoelastic characterization of tissue phantom, based on the photoacoustic measurement. The purpose of this study was to verify the applicability of the photoacoustic measurement method for monitoring of the development of extracellular matrix using tissue engineering technology. The decay times measured by the photoacoustic method were varied with culture periods when tissue-engineered articular cartilages with various culture periods (-12 weeks) were used as samples. Tissue-engineered cartilage cultured for a long period showed shorter decay times, indicating that the samples approached an elastic solid from a rheological viewpoint. By comparison between biochemical analyses and biomechanical studies, we proved that the photoacoustic signal was a good indicator for evaluating extracellular matrix formation because the change of the photoacoustic decay times would reflect the production of an extracellular matrix.

  9. Strategic Endothelial Cell Tube Formation Assay: Comparing Extracellular Matrix and Growth Factor Reduced Extracellular Matrix.

    PubMed

    Xie, Daniel; Ju, Donghong; Speyer, Cecilia; Gorski, David; Kosir, Mary A

    2016-01-01

    Malignant tumors require a blood supply in order to survive and spread. These tumors obtain their needed blood from the patient's blood stream by hijacking the process of angiogenesis, in which new blood vessels are formed from existing blood vessels. The CXCR2 (chemokine (C-X-C motif) receptor 2) receptor is a transmembrane G-protein-linked molecule found in many cells that is closely associated with angiogenesis(1). Specific blockade of the CXCR2 receptor inhibits angiogenesis, as measured by several assays such as the endothelial tube formation assay. The tube formation assay is useful for studying angiogenesis because it is an excellent method of studying the effects that any given compound or environmental condition may have on angiogenesis. It is a simple and quick in vitro assay that generates quantifiable data and requires relatively few components. Unlike in vivo assays, it does not require animals and can be carried out in less than two days. This protocol describes a variation of the extracellular matrix supporting endothelial tube formation assay, which tests the CXCR2 receptor. PMID:27585062

  10. Tailoring material properties of a nanofibrous extracellular matrix derived hydrogel

    NASA Astrophysics Data System (ADS)

    Johnson, Todd D.; Lin, Stephen Y.; Christman, Karen L.

    2011-12-01

    In the native tissue, the interaction between cells and the extracellular matrix (ECM) is essential for cell migration, proliferation, differentiation, mechanical stability, and signaling. It has been shown that decellularized ECMs can be processed into injectable formulations, thereby allowing for minimally invasive delivery. Upon injection and increase in temperature, these materials self-assemble into porous gels forming a complex network of fibers with nanoscale structure. In this study we aimed to examine and tailor the material properties of a self-assembling ECM hydrogel derived from porcine myocardial tissue, which was developed as a tissue specific injectable scaffold for cardiac tissue engineering. The impact of gelation parameters on ECM hydrogels has not previously been explored. We examined how modulating pH, temperature, ionic strength, and concentration affected the nanoscale architecture, mechanical properties, and gelation kinetics. These material characteristics were assessed using scanning electron microscopy, rheometry, and spectrophotometry, respectively. Since the main component of the myocardial matrix is collagen, many similarities between the ECM hydrogel and collagen gels were observed in terms of the nanofibrous structure and modulation of properties by altering ionic strength. However, variation from collagen gels was noted for the gelation temperature along with varied times and rates of gelation. These discrepancies when compared to collagen are likely due to the presence of other ECM components in the decellularized ECM based hydrogel. These results demonstrate how the material properties of ECM hydrogels could be tailored for future in vitro and in vivo applications.

  11. Extracellular Matrix Assembly in Diatoms (Bacillariophyceae)1

    PubMed Central

    Wustman, Brandon A.; Lind, Jan; Wetherbee, Richard; Gretz, Michael R.

    1998-01-01

    Achnanthes longipes is a marine, biofouling diatom that adheres to surfaces via adhesive polymers extruded during motility or organized into structures called stalks that contain three distinct regions: the pad, shaft, and collar. Four monoclonal antibodies (AL.C1–AL.C4) and antibodies from two uncloned hybridomas (AL.E1 and AL.E2) were raised against the extracellular adhesives of A. longipes. Antibodies were screened against a hot-water-insoluble/hot-bicarbonate-soluble-fraction. The hot-water-insoluble/hot-bicarbonate-soluble fraction was fractionated to yield polymers in three size ranges: F1, ≥ 20,000,000 Mr; F2, ≅100,000 Mr; and F3, <10,000 Mr relative to dextran standards. The ≅100,000-Mr fraction consisted of highly sulfated (approximately 11%) fucoglucuronogalactans (FGGs) and low-sulfate (approximately 2%) FGGs, whereas F1 was composed of O-linked FGG (F2)-polypeptide (F3) complexes. AL.C1, AL.C2, AL.C4, AL.E1, and AL.E2 recognized carbohydrate complementary regions on FGGs, with antigenicity dependent on fucosyl-containing side chains. AL.C3 was unique in that it had a lower affinity for FGGs and did not label any portion of the shaft. Enzyme-linked immunosorbent assay and immunocytochemistry indicated that low-sulfate FGGs are expelled from pores surrounding the raphe terminus, creating the cylindrical outer layers of the shaft, and that highly sulfated FGGs are extruded from the raphe, forming the central core. Antibody-labeling patterns and other evidence indicated that the shaft central-core region is related to material exuded from the raphe during cell motility. PMID:9536061

  12. Streptococcus mutans-derived extracellular matrix in cariogenic oral biofilms.

    PubMed

    Klein, Marlise I; Hwang, Geelsu; Santos, Paulo H S; Campanella, Osvaldo H; Koo, Hyun

    2015-01-01

    Biofilms are highly structured microbial communities that are enmeshed in a self-produced extracellular matrix. Within the complex oral microbiome, Streptococcus mutans is a major producer of extracellular polymeric substances including exopolysaccharides (EPS), eDNA, and lipoteichoic acid (LTA). EPS produced by S. mutans-derived exoenzymes promote local accumulation of microbes on the teeth, while forming a spatially heterogeneous and diffusion-limiting matrix that protects embedded bacteria. The EPS-rich matrix provides mechanical stability/cohesiveness and facilitates the creation of highly acidic microenvironments, which are critical for the pathogenesis of dental caries. In parallel, S. mutans also releases eDNA and LTA, which can contribute with matrix development. eDNA enhances EPS (glucan) synthesis locally, increasing the adhesion of S. mutans to saliva-coated apatitic surfaces and the assembly of highly cohesive biofilms. eDNA and other extracellular substances, acting in concert with EPS, may impact the functional properties of the matrix and the virulence of cariogenic biofilms. Enhanced understanding about the assembly principles of the matrix may lead to efficacious approaches to control biofilm-related diseases. PMID:25763359

  13. Streptococcus mutans-derived extracellular matrix in cariogenic oral biofilms

    PubMed Central

    Klein, Marlise I.; Hwang, Geelsu; Santos, Paulo H. S.; Campanella, Osvaldo H.; Koo, Hyun

    2015-01-01

    Biofilms are highly structured microbial communities that are enmeshed in a self-produced extracellular matrix. Within the complex oral microbiome, Streptococcus mutans is a major producer of extracellular polymeric substances including exopolysaccharides (EPS), eDNA, and lipoteichoic acid (LTA). EPS produced by S. mutans-derived exoenzymes promote local accumulation of microbes on the teeth, while forming a spatially heterogeneous and diffusion-limiting matrix that protects embedded bacteria. The EPS-rich matrix provides mechanical stability/cohesiveness and facilitates the creation of highly acidic microenvironments, which are critical for the pathogenesis of dental caries. In parallel, S. mutans also releases eDNA and LTA, which can contribute with matrix development. eDNA enhances EPS (glucan) synthesis locally, increasing the adhesion of S. mutans to saliva-coated apatitic surfaces and the assembly of highly cohesive biofilms. eDNA and other extracellular substances, acting in concert with EPS, may impact the functional properties of the matrix and the virulence of cariogenic biofilms. Enhanced understanding about the assembly principles of the matrix may lead to efficacious approaches to control biofilm-related diseases. PMID:25763359

  14. Rho GEFs and GAPs: Emerging integrators of extracellular matrix signaling

    PubMed Central

    Kutys, Matthew L; Yamada, Kenneth M

    2015-01-01

    Investigating cell migration in 3D settings has revealed that specific extracellular matrix environments require differential activities of the Rho GTPases for efficient migration. However, it is largely unknown how the activities of specific Rho GTPases are modulated to direct cell migration in response to different extracellular matrix cues. We have recently reported that extracellular matrix-dependent regulation of a specific Rho GEF is a fundamental mechanism governing cell migration in different microenvironments, providing a direct mechanism for extracellular matrix-specific regulation of Rho GTPase activity directing cell motility. We discovered that the Rho GEF βPix has a unique function during cell migration in fibrillar collagen environments by restraining RhoA signaling through a conserved signaling axis involving Cdc42 and the Rho GAP srGAP1. In this Commentary, we expand upon this new pathway and discuss potential mechanotransductive and therapeutic applications. Additionally, we speculate on a generalized role for Rho GEFs and GAPs in providing localized, context-dependent responses to the cellular microenvironment during cell migration and other cellular processes. PMID:25862162

  15. Specialisation of extracellular matrix for function in tendons and ligaments

    PubMed Central

    Birch, Helen L.; Thorpe, Chavaunne T.; Rumian, Adam P.

    2013-01-01

    Summary Tendons and ligaments are similar structures in terms of their composition, organisation and mechanical properties. The distinction between them stems from their anatomical location; tendons form a link between muscle and bone while ligaments link bones to bones. A range of overlapping functions can be assigned to tendon and ligaments and each structure has specific mechanical properties which appear to be suited for particular in vivo function. The extracellular matrix in tendon and ligament varies in accordance with function, providing appropriate mechanical properties. The most useful framework in which to consider extracellular matrix differences therefore is that of function rather than anatomical location. In this review we discuss what is known about the relationship between functional requirements, structural properties from molecular to gross level, cellular gene expression and matrix turnover. The relevance of this information is considered by reviewing clinical aspects of tendon and ligament repair and reconstructive procedures. PMID:23885341

  16. The Extracellular Matrix of Candida albicans Biofilms Impairs Formation of Neutrophil Extracellular Traps.

    PubMed

    Johnson, Chad J; Cabezas-Olcoz, Jonathan; Kernien, John F; Wang, Steven X; Beebe, David J; Huttenlocher, Anna; Ansari, Hamayail; Nett, Jeniel E

    2016-09-01

    Neutrophils release extracellular traps (NETs) in response to planktonic C. albicans. These complexes composed of DNA, histones, and proteins inhibit Candida growth and dissemination. Considering the resilience of Candida biofilms to host defenses, we examined the neutrophil response to C. albicans during biofilm growth. In contrast to planktonic C. albicans, biofilms triggered negligible release of NETs. Time lapse imaging confirmed the impairment in NET release and revealed neutrophils adhering to hyphae and migrating on the biofilm. NET inhibition depended on an intact extracellular biofilm matrix as physical or genetic disruption of this component resulted in NET release. Biofilm inhibition of NETosis could not be overcome by protein kinase C activation via phorbol myristate acetate (PMA) and was associated with suppression of neutrophil reactive oxygen species (ROS) production. The degree of impaired NET release correlated with resistance to neutrophil attack. The clinical relevance of the role for extracellular matrix in diminishing NET production was corroborated in vivo using a rat catheter model. The C. albicans pmr1Δ/Δ, defective in production of matrix mannan, appeared to elicit a greater abundance of NETs by scanning electron microscopy imaging, which correlated with a decreased fungal burden. Together, these findings show that C. albicans biofilms impair neutrophil response through an inhibitory pathway induced by the extracellular matrix. PMID:27622514

  17. Molecular Control of Vascular Tube Morphogenesis and Stabilization: Regulation by Extracellular Matrix, Matrix Metalloproteinases, and Endothelial Cell-Pericyte Interactions

    NASA Astrophysics Data System (ADS)

    Davis, George E.; Stratman, Amber N.; Sacharidou, Anastasia

    Recent studies have revealed a critical role for both extracellular matrices and matrix metalloproteinases in the molecular control of vascular morphogenesis and stabilization in three-dimensional (3D) tissue environments. Key interactions involve endothelial cells (ECs) and pericytes, which coassemble to affect vessel formation, remodeling, and stabilization events during development and postnatal life. EC-pericyte interactions control extracellular matrix remodeling events including vascular basement membrane matrix assembly, a necessary step for endothelial tube maturation and stabilization. ECs form tube networks in 3D extracellular matrices in a manner dependent on integrins, membrane-type metalloproteinases, and the Rho GTPases, Cdc42 and Rac1. Recent work has defined an EC lumen signaling complex of proteins composed of these proteins that controls 3D matrix-specific signaling events required for these processes. The EC tube formation process results in the creation of a network of proteolytically generated vascular guidance tunnels. These tunnels are physical matrix spaces that regulate vascular tube remodeling and represent matrix conduits into which pericytes are recruited to allow dynamic cell-cell interactions with ECs. These dynamic EC-pericyte interactions induce vascular basement membrane matrix deposition, leading to vessel maturation and stabilization.

  18. Extracellular matrix components direct porcine muscle stem cell behavior

    SciTech Connect

    Wilschut, Karlijn J.; Haagsman, Henk P.; Roelen, Bernard A.J.

    2010-02-01

    In muscle tissue, extracellular matrix proteins, together with the vasculature system, muscle-residence cells and muscle fibers, create the niche for muscle stem cells. The niche is important in controlling proliferation and directing differentiation of muscle stem cells to sustain muscle tissue. Mimicking the extracellular muscle environment improves tools exploring the behavior of primary muscle cells. Optimizing cell culture conditions to maintain muscle commitment is important in stem cell-based studies concerning toxicology screening, ex vivo skeletal muscle tissue engineering and in the enhancement of clinical efficiency. We used the muscle extracellular matrix proteins collagen type I, fibronectin, laminin, and also gelatin and Matrigel as surface coatings of tissue culture plastic to resemble the muscle extracellular matrix. Several important factors that determine myogenic commitment of the primary muscle cells were characterized by quantitative real-time RT-PCR and immunofluorescence. Adhesion of high PAX7 expressing satellite cells was improved if the cells were cultured on fibronectin or laminin coatings. Cells cultured on Matrigel and laminin coatings showed dominant integrin expression levels and exhibited an activated Wnt pathway. Under these conditions both stem cell proliferation and myogenic differentiation capacity were superior if compared to cells cultured on collagen type I, fibronectin and gelatin. In conclusion, Matrigel and laminin are the preferred coatings to sustain the proliferation and myogenic differentiation capacity of the primary porcine muscle stem cells, when cells are removed from their natural environment for in vitro culture.

  19. Extracellular Matrix and the Mechanics of Large Artery Development

    PubMed Central

    Cheng, Jeffrey K.; Wagenseil, Jessica E.

    2012-01-01

    The large, elastic arteries, as their name suggests, provide elastic distention and recoil during the cardiac cycle in vertebrate animals. The arteries are distended from the pressure of ejecting blood during active contraction of the left ventricle (LV) during systole, and recoil to their original dimensions during relaxation of the LV during diastole. The cyclic distension occurs with minimal energy loss, due to the elastic properties of one of the major structural extracellular matrix (ECM) components, elastin. The maximum distension is limited to prevent damage to the artery by another major ECM component, collagen. The mix of ECM components in the wall largely determines the passive mechanical behavior of the arteries and the subsequent load on the heart during systole. While much research has focused on initial artery formation, there has been less attention on the continuing development of the artery to produce the mature composite wall complete with endothelial cells (ECs), smooth muscle cells (SMCs), and the necessary mix of ECM components for proper cardiovascular function. This review focuses on the physiology of large artery development, including SMC differentiation and ECM production. The effects of hemodynamic forces and ECM deposition on the evolving arterial structure and function are discussed. Human diseases and mouse models with genetic mutations in ECM proteins that affect large artery development are summarized. A review of constitutive models and growth and remodeling theories is presented, along with future directions to improve understanding of ECM and the mechanics of large artery development. PMID:22584609

  20. Responses of fibroblasts to anchorage of dorsal extracellular matrix receptors.

    PubMed

    Beningo, Karen A; Dembo, Micah; Wang, Yu-li

    2004-12-28

    Fibroblasts in 2D cultures differ dramatically in behavior from those in the 3D environment of a multicellular organism. However, the basis of this disparity is unknown. A key difference is the spatial arrangement of anchored extracellular matrix (ECM) receptors to the ventral surface in 2D cultures and throughout the entire surface in 3D cultures. Therefore, we asked whether changing the topography of ECM receptor anchorage alone could invoke a morphological response. By using polyacrylamide-based substrates to present anchored fibronectin or collagen on dorsal cell surfaces, we found that well spread fibroblasts in 2D cultures quickly changed into a bipolar or stellate morphology similar to fibroblasts in vivo. Cells in this environment lacked lamellipodia and large actin bundles and formed small focal adhesions only near focused sites of protrusion. These responses depend on substrate rigidity, calcium ion, and, likely, the calcium-dependent protease calpain. We suggest that fibroblasts respond to both spatial distribution and mechanical input of anchored ECM receptors. Changes in cell shape may in turn affect diverse cellular activities, including gene expression, growth, and differentiation, as shown in numerous previous studies. PMID:15601776

  1. Targeting the neural extracellular matrix in neurological disorders.

    PubMed

    Soleman, S; Filippov, M A; Dityatev, A; Fawcett, J W

    2013-12-01

    The extracellular matrix (ECM) is known to regulate important processes in neuronal cell development, activity and growth. It is associated with the structural stabilization of neuronal processes and synaptic contacts during the maturation of the central nervous system. The remodeling of the ECM during both development and after central nervous system injury has been shown to affect neuronal guidance, synaptic plasticity and their regenerative responses. Particular interest has focused on the inhibitory role of chondroitin sulfate proteoglycans (CSPGs) and their formation into dense lattice-like structures, termed perineuronal nets (PNNs), which enwrap sub-populations of neurons and restrict plasticity. Recent studies in mammalian systems have implicated CSPGs and PNNs in regulating and restricting structural plasticity. The enzymatic degradation of CSPGs or destabilization of PNNs has been shown to enhance neuronal activity and plasticity after central nervous system injury. This review focuses on the role of the ECM, CSPGs and PNNs; and how developmental and pharmacological manipulation of these structures have enhanced neuronal plasticity and aided functional recovery in regeneration, stroke, and amblyopia. In addition to CSPGs, this review also points to the functions and potential therapeutic value of these and several other key ECM molecules in epileptogenesis and dementia. PMID:24012743

  2. Brain extracellular matrix retains connectivity in neuronal networks

    PubMed Central

    Bikbaev, Arthur; Frischknecht, Renato; Heine, Martin

    2015-01-01

    The formation and maintenance of connectivity are critically important for the processing and storage of information in neuronal networks. The brain extracellular matrix (ECM) appears during postnatal development and surrounds most neurons in the adult mammalian brain. Importantly, the removal of the ECM was shown to improve plasticity and post-traumatic recovery in the CNS, but little is known about the mechanisms. Here, we investigated the role of the ECM in the regulation of the network activity in dissociated hippocampal cultures grown on microelectrode arrays (MEAs). We found that enzymatic removal of the ECM in mature cultures led to transient enhancement of neuronal activity, but prevented disinhibition-induced hyperexcitability that was evident in age-matched control cultures with intact ECM. Furthermore, the ECM degradation followed by disinhibition strongly affected the network interaction so that it strongly resembled the juvenile pattern seen in naïve developing cultures. Taken together, our results demonstrate that the ECM plays an important role in retention of existing connectivity in mature neuronal networks that can be exerted through synaptic confinement of glutamate. On the other hand, removal of the ECM can play a permissive role in modification of connectivity and adaptive exploration of novel network architecture. PMID:26417723

  3. The Extracellular Matrix Contributes to Mechanotransduction in Uterine Fibroids

    PubMed Central

    Leppert, Phyllis C.; Jayes, Friederike L.; Segars, James H.

    2014-01-01

    The role of the extracellular matrix (ECM) and mechanotransduction as an important signaling factor in the human uterus is just beginning to be appreciated. The ECM is not only the substance that surrounds cells, but ECM stiffness will either compress cells or stretch them resulting in signals converted into chemical changes within the cell, depending on the amount of collagen, cross-linking, and hydration, as well as other ECM components. In this review we present evidence that the stiffness of fibroid tissue has a direct effect on the growth of the tumor through the induction of fibrosis. Fibrosis has two characteristics: (1) resistance to apoptosis leading to the persistence of cells and (2) secretion of collagen and other components of the ECM such a proteoglycans by those cells leading to abundant disposition of highly cross-linked, disoriented, and often widely dispersed collagen fibrils. Fibrosis affects cell growth by mechanotransduction, the dynamic signaling system whereby mechanical forces initiate chemical signaling in cells. Data indicate that the structurally disordered and abnormally formed ECM of uterine fibroids contributes to fibroid formation and growth. An appreciation of the critical role of ECM stiffness to fibroid growth may lead to new strategies for treatment of this common disease. PMID:25110476

  4. The Extracellular Matrix Regulates Granuloma Necrosis in Tuberculosis.

    PubMed

    Al Shammari, Basim; Shiomi, Takayuki; Tezera, Liku; Bielecka, Magdalena K; Workman, Victoria; Sathyamoorthy, Tarangini; Mauri, Francesco; Jayasinghe, Suwan N; Robertson, Brian D; D'Armiento, Jeanine; Friedland, Jon S; Elkington, Paul T

    2015-08-01

    A central tenet of tuberculosis pathogenesis is that caseous necrosis leads to extracellular matrix destruction and bacterial transmission. We reconsider the underlying mechanism of tuberculosis pathology and demonstrate that collagen destruction may be a critical initial event, causing caseous necrosis as opposed to resulting from it. In human tuberculosis granulomas, regions of extracellular matrix destruction map to areas of caseous necrosis. In mice, transgenic expression of human matrix metalloproteinase 1 causes caseous necrosis, the pathological hallmark of human tuberculosis. Collagen destruction is the principal pathological difference between humanised mice and wild-type mice with tuberculosis, whereas the release of proinflammatory cytokines does not differ, demonstrating that collagen breakdown may lead to cell death and caseation. To investigate this hypothesis, we developed a 3-dimensional cell culture model of tuberculosis granuloma formation, using bioelectrospray technology. Collagen improved survival of Mycobacterium tuberculosis-infected cells analyzed on the basis of a lactate dehydrogenase release assay, propidium iodide staining, and measurement of the total number of viable cells. Taken together, these findings suggest that collagen destruction is an initial event in tuberculosis immunopathology, leading to caseous necrosis and compromising the immune response, revealing a previously unappreciated role for the extracellular matrix in regulating the host-pathogen interaction. PMID:25676469

  5. Vascular wall extracellular matrix proteins and vascular diseases

    PubMed Central

    Xu, Junyan; Shi, Guo-Ping

    2014-01-01

    Extracellular matrix proteins form the basic structure of blood vessels. Along with providing basic structural support to blood vessels, matrix proteins interact with different sets of vascular cells via cell surface integrin or non-integrin receptors. Such interactions induce vascular cell de novo synthesis of new matrix proteins during blood vessel development or remodeling. Under pathological conditions, vascular matrix proteins undergo proteolytic processing, yielding bioactive fragments to influence vascular wall matrix remodeling. Vascular cells also produce alternatively spliced variants that induce vascular cell production of different matrix proteins to interrupt matrix homeostasis, leading to increased blood vessel stiffness; vascular cell migration, proliferation, or death; or vascular wall leakage and rupture. Destruction of vascular matrix proteins leads to vascular cell or blood-borne leukocyte accumulation, proliferation, and neointima formation within the vascular wall; blood vessels prone to uncontrolled enlargement during blood flow diastole; tortuous vein development; and neovascularization from existing pathological tissue microvessels. Here we summarize discoveries related to blood vessel matrix proteins within the past decade from basic and clinical studies in humans and animals — from expression to cross-linking, assembly, and degradation under physiological and vascular pathological conditions, including atherosclerosis, aortic aneurysms, varicose veins, and hypertension. PMID:25045854

  6. Gene evolution and functions of extracellular matrix proteins in teeth.

    PubMed

    Yoshizaki, Keigo; Yamada, Yoshihiko

    2013-03-01

    The extracellular matrix (ECM) not only provides physical support for tissues, but it is also critical for tissue development, homeostasis and disease. Over 300 ECM molecules have been defined as comprising the "core matrisome" in mammals through the analysis of whole genome sequences. During tooth development, the structure and functions of the ECM dynamically change. In the early stages, basement membranes (BMs) separate two cell layers of the dental epithelium and the mesenchyme. Later in the differentiation stages, the BM layer is replaced with the enamel matrix and the dentin matrix, which are secreted by ameloblasts and odontoblasts, respectively. The enamel matrix genes and the dentin matrix genes are each clustered in two closed regions located on human chromosome 4 (mouse chromosome 5), except for the gene coded for amelogenin, the major enamel matrix protein, which is located on the sex chromosomes. These genes for enamel and dentin matrix proteins are derived from a common ancestral gene, but as a result of evolution, they diverged in terms of their specific functions. These matrix proteins play important roles in cell adhesion, polarity, and differentiation and mineralization of enamel and dentin matrices. Mutations of these genes cause diseases such as odontogenesis imperfect (OI) and amelogenesis imperfect (AI). In this review, we discuss the recently defined terms matrisome and matrixome for ECMs, as well as focus on genes and functions of enamel and dentin matrix proteins. PMID:23539364

  7. [The corneal wound healing and the extracellular matrix].

    PubMed

    Varkoly, Gréta; Bencze, János; Hortobágyi, Tibor; Módis, László

    2016-06-19

    The cornea is the first refractive element of the eye. The transparency of the cornea results from the regularly arranged collagen fibrils, forming lamellar structure and the leucin rich proteoglycans, which make interactions between the fibrils. The adult cornea consists mainly of fibril-forming collagens. The cornea has less amount of fibril associated and non-fibrillar collagens. The main proteoglycans of the cornea are keratan-sulfate proteoglycans and it also contains dermatan-sulfate proteoglycans. Disorders of the proteoglycan synthesis lead to the disruption of the unique pattern and result in thicker collagen fibrils. The abnormal structure of the extracellular matrix can generate corneal disorders and the loss of corneal transparency. Furthermore, proteoglycans and collagens have an important role in wound healing. In injury the keratocytes produce higher amounts of collagens and proteoglycans mediated by growth factors. Depending on the ratio of the cells and growth factors the extracellular matrix returns to normal or corneal scar tissue develops. PMID:27287839

  8. Conformal Nanopatterning of Extracellular Matrix Proteins onto Topographically Complex Surfaces

    PubMed Central

    Sun, Yan; Jallerat, Quentin; Szymanski, John M.

    2015-01-01

    We report a method for conformal nanopatterning of extracellular matrix proteins onto engineered surfaces independent of underlying microtopography. This enables fibronectin, laminin, and other proteins to be applied to biomaterial surfaces in complex geometries inaccessible using traditional soft lithography techniques. Engineering combinatorial surfaces that integrate topographical and biochemical micropatterns enhances control of the biotic-abiotic interface, used here to understand cardiomyocyte response to competing physical and chemical cues in the microenvironment. PMID:25506720

  9. Fibulin's organization into the extracellular matrix of fetal lung fibroblasts is dependent on fibronectin matrix assembly.

    PubMed

    Roman, J; McDonald, J A

    1993-05-01

    Fibulin is a newly described extracellular matrix (ECM) glycoprotein whose function has not been elucidated. We have observed that cultured fetal lung fibroblasts produce fibulin and have postulated that its expression may be important during lung development. To begin to understand the potential function of fibulin in lung development, we examined its expression and distribution in cultured fetal lung fibroblasts. Immunofluorescence staining of cultured fibroblasts revealed that fibulin was distributed upon their surface in a fibrillar array resembling fibronectin (FN), another ECM glycoprotein expressed by fetal lung fibroblasts and implicated in lung and heart development. Detection of fibulin by immunofluorescence staining of nonpermeabilized cells, its immunoprecipitation from 125I-cell surface-labeled fibroblasts, pulse-chase analysis, and temperature-induced phase separation studies revealed that fibulin is an ECM peripheral membrane protein that is synthesized and secreted by cultured fetal lung fibroblasts shortly after plating and incorporated into their matrix in a divalent cation-dependent manner. Because fibulin co-distributes with both FN and the FN receptor, the integrin alpha 5 beta 1, we examined the possibility that fibulin was interacting with both components. Dissociation of FN receptors from FN fibers with anti-FN receptor antibodies did not affect fibulin's distribution, suggesting that fibulin binds FN and that this interaction is not affected by the state of FN receptor binding. Finally, inhibition of FN matrix assembly prevented the deposition of fibulin, providing further support for FN-fibulin interactions and suggesting that fibulin deposition is dependent on FN matrix assembly. PMID:8481235

  10. Fibronectin Deposition Participates in Extracellular Matrix Assembly and Vascular Morphogenesis

    PubMed Central

    Hielscher, Abigail; Ellis, Kim; Qiu, Connie; Porterfield, Josh; Gerecht, Sharon

    2016-01-01

    The extracellular matrix (ECM) has been demonstrated to facilitate angiogenesis. In particular, fibronectin has been documented to activate endothelial cells, resulting in their transition from a quiescent state to an active state in which the cells exhibit enhanced migration and proliferation. The goal of this study is to examine the role of polymerized fibronectin during vascular tubulogenesis using a 3 dimensional (3D) cell-derived de-cellularized matrix. A fibronectin-rich 3D de-cellularized ECM was used as a scaffold to study vascular morphogenesis of endothelial cells (ECs). Confocal analyses of several matrix proteins reveal high intra- and extra-cellular deposition of fibronectin in formed vascular structures. Using a small peptide inhibitor of fibronectin polymerization, we demonstrate that inhibition of fibronectin fibrillogenesis in ECs cultured atop de-cellularized ECM resulted in decreased vascular morphogenesis. Further, immunofluorescence and ultrastructural analyses reveal decreased expression of stromal matrix proteins in the absence of polymerized fibronectin with high co-localization of matrix proteins found in association with polymerized fibronectin. Evaluating vascular kinetics, live cell imaging showed that migration, migration velocity, and mean square displacement, are disrupted in structures grown in the absence of polymerized fibronectin. Additionally, vascular organization failed to occur in the absence of a polymerized fibronectin matrix. Consistent with these observations, we tested vascular morphogenesis following the disruption of EC adhesion to polymerized fibronectin, demonstrating that block of integrins α5β1 and αvβ3, abrogated vascular morphogenesis. Overall, fibronectin deposition in a 3D cell-derived de-cellularized ECM appears to be imperative for matrix assembly and vascular morphogenesis. PMID:26811931

  11. Degradation of extracellular matrix and its components by hypobromous acid

    PubMed Central

    Rees, Martin D.; McNiven, Tane N.; Davies, Michael J.

    2006-01-01

    EPO (eosinophil peroxidase) and MPO (myeloperoxidase) are highly basic haem enzymes that can catalyse the production of HOBr (hypobromous acid). They are released extracellularly by activated leucocytes and their binding to the polyanionic glycosa-minoglycan components of extracellular matrix (proteoglycans and hyaluronan) may localize the production of HOBr to these materials. It is shown in the present paper that the reaction of HOBr with glycosaminoglycans (heparan sulfate, heparin, chondroitin sulfate and hyaluronan) generates polymer-derived N-bromo derivatives (bromamines, dibromamines, N-bromosulfon-amides and bromamides). Decomposition of these species, which can occur spontaneously and/or via one-electron reduction by low-valent transition metal ions (Cu+ and Fe2+), results in polymer fragmentation and modification. One-electron reduction of the N-bromo derivatives generates radicals that have been detected by EPR spin trapping. The species detected are consistent with metal ion-dependent polymer fragmentation and modification being initiated by the formation of nitrogen-centred (aminyl, N-bromoaminyl, sulfonamidyl and amidyl) radicals. Previous studies have shown that the reaction of HOBr with proteins generates N-bromo derivatives and results in fragmentation of the polypeptide backbone. The reaction of HOBr with extracellular matrix synthesized by smooth muscle cells in vitro induces the release of carbohydrate and protein components in a time-dependent manner, which is consistent with fragmentation of these materials via the formation of N-bromo derivatives. The degradation of extracellular matrix glycosaminoglycans and proteins by HOBr may contribute to tissue damage associated with inflammatory diseases such as asthma. PMID:17014424

  12. Macromolecular Crowding Directs Extracellular Matrix Organization and Mesenchymal Stem Cell Behavior

    PubMed Central

    Zeiger, Adam S.; Loe, Felicia C.; Li, Ran; Raghunath, Michael; Van Vliet, Krystyn J.

    2012-01-01

    Microenvironments of biological cells are dominated in vivo by macromolecular crowding and resultant excluded volume effects. This feature is absent in dilute in vitro cell culture. Here, we induced macromolecular crowding in vitro by using synthetic macromolecular globules of nm-scale radius at physiological levels of fractional volume occupancy. We quantified the impact of induced crowding on the extracellular and intracellular protein organization of human mesenchymal stem cells (MSCs) via immunocytochemistry, atomic force microscopy (AFM), and AFM-enabled nanoindentation. Macromolecular crowding in extracellular culture media directly induced supramolecular assembly and alignment of extracellular matrix proteins deposited by cells, which in turn increased alignment of the intracellular actin cytoskeleton. The resulting cell-matrix reciprocity further affected adhesion, proliferation, and migration behavior of MSCs. Macromolecular crowding can thus aid the design of more physiologically relevant in vitro studies and devices for MSCs and other cells, by increasing the fidelity between materials synthesized by cells in vivo and in vitro. PMID:22649562

  13. Milk extracellular vesicles accelerate osteoblastogenesis but impair bone matrix formation.

    PubMed

    Oliveira, Marina C; Arntz, Onno J; Blaney Davidson, Esmeralda N; van Lent, Peter L E M; Koenders, Marije I; van der Kraan, Peter M; van den Berg, Wim B; Ferreira, Adaliene V M; van de Loo, Fons A J

    2016-04-01

    The claimed beneficial effect of milk on bone is still a matter for debate. Recently extracellular vesicles (EVs) that contain proteins and RNA were discovered in milk, but their effect on bone formation has not yet been determined. We demonstrated previously that bovine milk-derived EVs (BMEVs) have immunoregulatory properties. Our aim was to evaluate the effect of BMEVs on osteogenesis by mice and human mesenchymal stem cells (hMSCs). Oral delivery of two concentrations of BMEVs to female DBA/1J mice during 7weeks did not alter the tibia trabecular bone area; however, the osteocytes number increased. In addition, the highest dose of BMEVs markedly increased the woven bone tissue, which is more brittle. The exposure of hMSCs to BMEVs during 21days resulted in less mineralization but higher cell proliferation. Interestingly BMEVs reduced the collagen production, but enhanced the expression of genes characteristic for immature osteoblasts. A kinetic study showed that BMEVs up-regulated many osteogenic genes within the first 4days. However, the production of type I collagen and expression of its genes (COL1A1 and COL1A2) were markedly reduced at days 21 and 28. At day 28, BMEVs again lead to higher proliferation, but mineralization was significantly increased. This was associated with increased expression of sclerostin, a marker for osteocytes, and reduced osteonectin, which is associated to bone matrix formation. Our study adds BMEVs to the list of milk components that can affect bone formation and may shed new light on the contradictory claims of milk on bone formation. PMID:27012623

  14. Structural Determinants of Binding of Aromates to Extracellular Matrix

    PubMed Central

    Zhang, Yufen; Lukacova, Viera; Bartus, Vladimir; Balaz, Stefan

    2008-01-01

    For small molecules acting in tissues, including signaling peptides, effectors, inhibitors, and other drug candidates, nonspecific binding to the extracellular matrix (ECM) is a critical phenomenon affecting their disposition, toxicity, and other effects. A commercially available ECM mimic, forming a solidified layer at the bottom of the vials, was used to measure the association constants of 25 simple aromatic compounds to two forms of ECM proteins, solidified (s-ECM) and dissolved (d-ECM) in the buffer during the incubation. Except for small homologous series, the binding data did not correlate with the lipophilicity and acidity of the compounds, contrary to a common expectation for nonspecific binding. To elucidate the putative structures of averaged binding sites of s-ECM and d-ECM, the Comparative Molecular Field Analysis (CoMFA) was applied in a modified version taking into consideration that multiple modes and multiple species may be involved. The method shapes a receptor site model from a set of grid points, in which the interaction energies between a probe atom and superimposed ligands are calculated. Electrostatic and steric energies in the grid points are characterized by regression coefficients. The forward-selection, nonlinear regression analysis was used to optimize the coefficients in the novel multi-species, multi-mode CoMFA models. These models showed satisfactory descriptive and predictive abilities for both s-ECM and d-ECM binding data, which were better than those obtained with the standard, one-mode CoMFA analysis. The calibrated models, defining the electrostatic and van der Waals regions of putative binding sites, are suitable for the prediction of ECM binding for untested chemicals. PMID:17226922

  15. The extracellular matrix as a modulator of the inflammatory and reparative response following myocardial infarction

    PubMed Central

    Dobaczewski, Marcin; Gonzalez-Quesada, Carlos; Frangogiannis, Nikolaos G

    2009-01-01

    The dynamic alterations in the cardiac extracellular matrix following myocardial infarction not only determine the mechanical properties of the infarcted heart, but also directly modulate the inflammatory and reparative response. During the inflammatory phase of healing, rapid activation of matrix metalloproteinases (MMP) causes degradation of the cardiac extracellular matrix. Matrix fragments exert potent pro-inflammatory actions, while MMPs process cytokines and chemokines altering their biological activity. In addition, vascular hyperpermeability results in extravasation of fibronectin and fibrinogen leading to formation of a plasma-derived provisional matrix that serves as a scaffold for leukocyte infiltration. Clearance of the infarct from dead cells and matrix debris is essential for resolution of inflammation and marks the transition to the proliferative phase. The fibrin-based provisional matrix is lysed and cellular fibronectin is secreted. ED-A fibronectin, mechanical tension and Transforming Growth Factor (TGF)-β are essential for modulation of fibroblasts into myofibroblasts, the main collagen-secreting cells in the wound. The matricellular proteins thrombospondin-1 and -2, osteopontin, tenascin-C, periostin, and secreted protein acidic and rich in cysteine (SPARC) are induced in the infarct regulating cellular interactions and promoting matrix organization. As the infarct matures, matrix cross-linking results in formation of a dense collagen-based scar. At this stage, shielding of fibroblasts from external mechanical tension by the mature matrix network may promote deactivation and cellular quiescence. The components of the extracellular matrix do not passively follow the pathologic alterations of the infarcted heart but critically modulate inflammatory and reparative pathways by transducing signals that affect cell survival, phenotype and gene expression. PMID:19631653

  16. Astrocytes and extracellular matrix in extrasynaptic volume transmission

    PubMed Central

    Vargová, Lýdia; Syková, Eva

    2014-01-01

    Volume transmission is a form of intercellular communication that does not require synapses; it is based on the diffusion of neuroactive substances across the brain extracellular space (ECS) and their binding to extrasynaptic high-affinity receptors on neurons or glia. Extracellular diffusion is restricted by the limited volume of the ECS, which is described by the ECS volume fraction α, and the presence of diffusion barriers, reflected by tortuosity λ, that are created, for example, by fine astrocytic processes or extracellular matrix (ECM) molecules. Organized astrocytic processes, ECM scaffolds or myelin sheets channel the extracellular diffusion so that it is facilitated in a certain direction, i.e. anisotropic. The diffusion properties of the ECS are profoundly influenced by various processes such as the swelling and morphological rebuilding of astrocytes during either transient or persisting physiological or pathological states, or the remodelling of the ECM in tumorous or epileptogenic tissue, during Alzheimer's disease, after enzymatic treatment or in transgenic animals. The changing diffusion properties of the ECM influence neuron–glia interaction, learning abilities, the extent of neuronal damage and even cell migration. From a clinical point of view, diffusion parameter changes occurring during pathological states could be important for diagnosis, drug delivery and treatment. PMID:25225101

  17. Structure and Function of the Skeletal Muscle Extracellular Matrix

    PubMed Central

    Gillies, Allison R.; Lieber, Richard L.

    2011-01-01

    The skeletal muscle extracellular matrix (ECM) plays an important role in muscle fiber force transmission, maintenance, and repair. In both injured and diseased states, ECM adapts dramatically, a property thathas clinical manifestations and alters muscle function. Here, we review the structure, composition, and mechanical properties of skeletal muscle ECM, describe the cells that contribute to the maintenance of the ECM and, finally, overview changes that occur with pathology. New scanning electron micrographs of ECM structure are also presented with hypotheses about ECM structure-function relationships. Detailed structure-function relationships of the ECM have yet to be defined and, as a result, we propose areas for future studies. PMID:21949456

  18. Rat pancreatic stellate cells secrete matrix metalloproteinases: implications for extracellular matrix turnover

    PubMed Central

    Phillips, P A; McCarroll, J A; Park, S; Wu, M-J; Pirola, R; Korsten, M; Wilson, J S; Apte, M V

    2003-01-01

    weights consistent with MMP2, MMP9, and MMP13. RT-PCR demonstrated the presence of mRNA for metalloproteinase inhibitors TIMP1 and TIMP2 in PSCs while reverse zymography revealed the presence of functional TIMP2 in PSC secretions. MMP2 secretion by PSCs was significantly increased by TGF-β1 and IL-6, but was not affected by TNF-α. Ethanol and acetaldehyde induced secretion of both MMP2 and TIMP2 by PSCs. Conclusions: Pancreatic stellate cells have the capacity to synthesise a number of matrix metalloproteinases, including MMP2, MMP9, and MMP13 and their inhibitors TIMP1 and TIMP2. MMP2 secretion by PSCs is significantly increased on exposure to the proinflammatory cytokines TGF-β1 and IL-6. Both ethanol and its metabolite acetaldehyde increase MMP2 as well as TIMP2 secretion by PSCs. Implication: The role of pancreatic stellate cells in extracellular matrix formation and fibrogenesis may be related to their capacity to regulate the degradation as well as the synthesis of extracellular matrix proteins. PMID:12524413

  19. Streptococcus pyogenes degrades extracellular matrix in chondrocytes via MMP-13

    SciTech Connect

    Sakurai, Atsuo; Okahashi, Nobuo; Maruyama, Fumito; Ooshima, Takashi; Hamada, Shigeyuki; Nakagawa, Ichiro

    2008-08-29

    Group A streptococcus (GAS) causes a wide range of human diseases, including bacterial arthritis. The pathogenesis of arthritis is characterized by synovial proliferation and the destruction of cartilage and subchondral bone in joints. We report here that GAS strain JRS4 invaded a chondrogenic cell line ATDC5 and induced the degradation of the extracellular matrix (ECM), whereas an isogenic mutant of JRS4 lacking a fibronectin-binding protein, SAM1, failed to invade the chondrocytes or degrade the ECM. Reverse transcription-PCR and Western blot analysis revealed that the expression of matrix metalloproteinase (MMP)-13 was strongly elevated during the infection with GAS. A reporter assay revealed that the activation of the AP-1 transcription factor and the phosphorylation of c-Jun terminal kinase participated in MMP-13 expression. These results suggest that MMP-13 plays an important role in the destruction of infected joints during the development of septic arthritis.

  20. The design of reversible hydrogels to capture extracellular matrix dynamics

    NASA Astrophysics Data System (ADS)

    Rosales, Adrianne M.; Anseth, Kristi S.

    2016-02-01

    The extracellular matrix (ECM) is a dynamic environment that constantly provides physical and chemical cues to embedded cells. Much progress has been made in engineering hydrogels that can mimic the ECM, but hydrogel properties are, in general, static. To recapitulate the dynamic nature of the ECM, many reversible chemistries have been incorporated into hydrogels to regulate cell spreading, biochemical ligand presentation and matrix mechanics. For example, emerging trends include the use of molecular photoswitches or biomolecule hybridization to control polymer chain conformation, thereby enabling the modulation of the hydrogel between two states on demand. In addition, many non-covalent, dynamic chemical bonds have found increasing use as hydrogel crosslinkers or tethers for cell signalling molecules. These reversible chemistries will provide greater temporal control of adhered cell behaviour, and they allow for more advanced in vitro models and tissue-engineering scaffolds to direct cell fate.

  1. Extracellular matrix and its receptors in Drosophila neural development

    PubMed Central

    Broadie, Kendal; Baumgartner, Stefan; Prokop, Andreas

    2011-01-01

    Extracellular matrix (ECM) and matrix receptors are intimately involved in most biological processes. The ECM plays fundamental developmental and physiological roles in health and disease, including processes underlying the development, maintenance and regeneration of the nervous system. To understand the principles of ECM-mediated functions in the nervous system, genetic model organisms like Drosophila provide simple, malleable and powerful experimental platforms. This article provides an overview of ECM proteins and receptors in Drosophila. It then focuses on their roles during three progressive phases of neural development: 1) neural progenitor proliferation, 2) axonal growth and pathfinding and 3) synapse formation and function. Each section highlights known ECM and ECM-receptor components and recent studies done in mutant conditions to reveal their in vivo functions, all illustrating the enormous opportunities provided when merging work on the nervous system with systematic research into ECM-related gene functions. PMID:21688401

  2. The Extracellular Matrix In Development and Morphogenesis: A Dynamic View

    PubMed Central

    Rozario, Tania; DeSimone, Douglas W.

    2009-01-01

    The extracellular matrix (ECM) is synthesized and secreted by embryonic cells beginning at the earliest stages of development. Our understanding of ECM composition, structure and function has grown considerably in the last several decades and this knowledge has revealed that the extracellular microenvironment is critically important for cell growth, survival, differentiation and morphogenesis. ECM and the cellular receptors that interact with it mediate both physical linkages with the cytoskeleton and the bidirectional flow of information between the extracellular and intracellular compartments. This review considers the range of cell and tissue functions attributed to ECM molecules and summarizes recent findings specific to key developmental processes. The importance of ECM as a dynamic repository for growth factors is highlighted along with more recent studies implicating the 3-dimensional organization and physical properties of the ECM as it relates to cell signaling and the regulation of morphogenetic cell behaviors. Embryonic cell and tissue generated forces and mechanical signals arising from ECM adhesion represent emerging areas of interest in this field. PMID:19854168

  3. Extracellular matrix of the bovine ovarian membrana granulosa.

    PubMed

    Rodgers, R J; Irving Rodgers, H F

    2002-05-31

    Much is known about the control of the development of ovarian follicles by growth factors and hormones. The study of extracellular matrix in the ovary, though, is a relatively new area. To date much research has focused on identifying the matrix components present, and more recently, its production and the physiological roles. In this review we focus on the changes that occur in the follicular basal lamina from primordial follicles through to ovulation and formation of the corpus luteum, the changes that occur during follicular atresia, and we discuss our observations of a novel matrix which forms in the membrana granulosa. The follicular basal lamina changes considerably during follicular development in its expression pattern of type IV collagens. Of the laminin chains examined, there appears only to be an increase in amount, except for laminin alpha2. It is expressed only in a small proportion of healthy antral follicles and in the majority of atretic antral follicles. Call-Exner bodies have the same composition as the basal lamina, except they do not contain laminin alpha2, even when the follicular basal lamina does. The novel matrix that develops within the membrana granulosa is similar in composition to Call-Exner bodies which occur predominantly in preantral follicles, except that it is far more common in large antral follicles, does not induce polarization of the surrounding granulosa cells, and does not contain follicular fluid-like material as the Call-Exner bodies of some species do. The expression of this matrix occurs prior to and during the time when granulosa cells express steroidogenic enzymes. It does not exist in corpora lutea. In addition large luteal cells, derived from granulosa cells, do not appear to have a basal lamina. These findings suggest that the maturational changes in the membrana granulosa are accompanied by changes in the matrix. PMID:12044919

  4. The Extracellular Matrix in Photosynthetic Mats: A Cyanobacterial Gingerbread House

    NASA Astrophysics Data System (ADS)

    Stuart, R.; Stannard, W.; Bebout, B.; Pett-Ridge, J.; Mayali, X.; Weber, P. K.; Lipton, M. S.; Lee, J.; Everroad, R. C.; Thelen, M.

    2014-12-01

    Hypersaline laminated cyanobacterial mats are excellent model systems for investigating photoautotrophic contributions to biogeochemical cycling on a millimeter scale. These self-sustaining ecosystems are characterized by steep physiochemical gradients that fluctuate dramatically on hour timescales, providing a dynamic environment to study microbial response. However, elucidating the distribution of energy from light absorption into biomass requires a complete understanding of the various constituents of the mat. Extracellular polymeric substances (EPS), which can be composed of proteins, polysaccharides, lipids and DNA are a major component of these mats and may function in the redistribution of nutrients and metabolites within the community. To test this notion, we established a model mat-building culture for comparison with the phylogenetically diverse natural mat communities. In these two systems we determined how proteins and glycans in the matrix changed as a function of light and tracked nutrient flow from the matrix. Using mass spectrometry metaproteomics analysis, we found homologous proteins in both field and culture extracellular matrix that point to cyanobacterial turnover of amino acids, inorganic nutrients, carbohydrates and nucleic acids from the EPS. Other abundant functions identified included oxidative stress response from both the cyanobacteria and heterotrophs and cyanobacterial structural proteins that may play a role in mat cohesion. Several degradative enzymes also varied in abundance in the EPS in response to light availability, suggesting active secretion. To further test cyanobacterial EPS turnover, we generated isotopically-labeled EPS and used NanoSIMS to trace uptake of this labeled EPS. Our findings suggest Cyanobacteria may facilitate nutrient transfer to other groups, as well as uptake of their own products through degradation of EPS components. This work provides evidence for the essential roles of EPS for storage, structural

  5. Synthetic osteogenic extracellular matrix formed by coated silicon dioxide nanosprings

    PubMed Central

    2012-01-01

    Background The design of biomimetic materials that parallel the morphology and biology of extracellular matrixes is key to the ability to grow functional tissues in vitro and to enhance the integration of biomaterial implants into existing tissues in vivo. Special attention has been put into mimicking the nanostructures of the extracellular matrix of bone, as there is a need to find biomaterials that can enhance the bonding between orthopedic devices and this tissue. Methods We have tested the ability of normal human osteoblasts to propagate and differentiate on silicon dioxide nanosprings, which can be easily grown on practically any surface. In addition, we tested different metals and metal alloys as coats for the nanosprings in tissue culture experiments with bone cells. Results Normal human osteoblasts grown on coated nanosprings exhibited an enhanced rate of propagation, differentiation into bone forming cells and mineralization. While osteoblasts did not attach effectively to bare nanowires grown on glass, these cells propagated successfully on nanosprings coated with titanium oxide and gold. We observed a 270 fold increase in the division rate of osteoblasts when grow on titanium/gold coated nanosprings. This effect was shown to be dependent on the nanosprings, as the coating by themselves did not alter the growth rate of osteoblast. We also observed that titanium/zinc/gold coated nanosprings increased the levels of osteoblast production of alkaline phosphatase seven folds. This result indicates that osteoblasts grown on this metal alloy coated nanosprings are differentiating to mature bone making cells. Consistent with this hypothesis, we showed that osteoblasts grown on the same metal alloy coated nanosprings have an enhanced ability to deposit calcium salt. Conclusion We have established that metal/metal alloy coated silicon dioxide nanosprings can be used as a biomimetic material paralleling the morphology and biology of osteogenic extracellular matrix

  6. Biomechanical and biochemical remodeling of stromal extracellular matrix in cancer.

    PubMed

    Malik, Ruchi; Lelkes, Peter I; Cukierman, Edna

    2015-04-01

    The extracellular matrix (ECM) provides structural and biochemical signals that regulate cell function. A well-controlled balance between cells and surroundings (i.e., dynamic reciprocity) is crucial for regulating ECM architecture. During cancer progression, epithelial cells undergo genetic alterations which, together with stromal changes including ECM remodeling, disturb the homeostatic dynamics of the epithelium. A parallel organization of stromal ECM fibrils is associated with tumorigenic responses. In an emerging paradigm, continuous and progressive regulation via mechanical forces and aberrant signaling are believed to be responsible for tumor-associated ECM remodeling. In this review we discuss the discrete biomechanical and biochemical mechanisms that underlie these architectural changes and highlight their particular relevance to the regulation of the alignment of ECM in the mesenchymal stroma. PMID:25708906

  7. Cell stiffness, contractile stress and the role of extracellular matrix

    SciTech Connect

    An, Steven S.; Kim, Jina; Ahn, Kwangmi; Trepat, Xavier; Drake, Kenneth J.; Kumar, Sarvesh; Ling, Guoyu; Purington, Carolyn; Rangasamy, Tirumalai; Kensler, Thomas W.; Mitzner, Wayne; Fredberg, Jeffrey J.; Biswal, Shyam

    2009-05-15

    Here we have assessed the effects of extracellular matrix (ECM) composition and rigidity on mechanical properties of the human airway smooth muscle (ASM) cell. Cell stiffness and contractile stress showed appreciable changes from the most relaxed state to the most contracted state: we refer to the maximal range of these changes as the cell contractile scope. The contractile scope was least when the cell was adherent upon collagen V, followed by collagen IV, laminin, and collagen I, and greatest for fibronectin. Regardless of ECM composition, upon adherence to increasingly rigid substrates, the ASM cell positively regulated expression of antioxidant genes in the glutathione pathway and heme oxygenase, and disruption of a redox-sensitive transcription factor, nuclear erythroid 2 p45-related factor (Nrf2), culminated in greater contractile scope. These findings provide biophysical evidence that ECM differentially modulates muscle contractility and, for the first time, demonstrate a link between muscle contractility and Nrf2-directed responses.

  8. Constructive remodeling of a synthetic endothelial extracellular matrix

    PubMed Central

    Han, Sewoon; Shin, Yoojin; Jeong, Hyo Eun; Jeon, Jessie S.; Kamm, Roger D.; Huh, Dongeun; Sohn, Lydia L.; Chung, Seok

    2015-01-01

    The construction of well-controllable in vitro models of physiological and pathological vascular endothelium remains a fundamental challenge in tissue engineering and drug development. Here, we present an approach for forming a synthetic endothelial extracellular matrix (ECM) that closely resembles that of the native structure by locally depositing basement membrane materials onto type 1 collagen nanofibers only in a region adjacent to the endothelial cell (EC) monolayer. Culturing the EC monolayer on this synthetic endothelial ECM remarkably enhanced its physiological properties, reducing its vascular permeability, and promoting a stabilized, quiescent phenotype. We demonstrated that the EC monolayer on the synthetic endothelial ECM neither creates non-physiological barriers to cell-cell or cell-ECM interactions, nor hinders molecular diffusion of growth factors and other molecules. The synthetic endothelial ECM and vascular endothelium on it may help us enter in a new phase of research in which various models of the biological barrier behavior can be tested experimentally. PMID:26687334

  9. Insight On Colorectal Carcinoma Infiltration by Studying Perilesional Extracellular Matrix

    PubMed Central

    Nebuloni, Manuela; Albarello, Luca; Andolfo, Annapaola; Magagnotti, Cinzia; Genovese, Luca; Locatelli, Irene; Tonon, Giovanni; Longhi, Erika; Zerbi, Pietro; Allevi, Raffaele; Podestà, Alessandro; Puricelli, Luca; Milani, Paolo; Soldarini, Armando; Salonia, Andrea; Alfano, Massimo

    2016-01-01

    The extracellular matrix (ECM) from perilesional and colorectal carcinoma (CRC), but not healthy colon, sustains proliferation and invasion of tumor cells. We investigated the biochemical and physical diversity of ECM in pair-wised comparisons of healthy, perilesional and CRC specimens. Progressive linearization and degree of organization of fibrils was observed from healthy to perilesional and CRC ECM, and was associated with a steady increase of stiffness and collagen crosslinking. In the perilesional ECM these modifications coincided with increased vascularization, whereas in the neoplastic ECM they were associated with altered modulation of matrisome proteins, increased content of hydroxylated lysine and lysyl oxidase. This study identifies the increased stiffness and crosslinking of the perilesional ECM predisposing an environment suitable for CRC invasion as a phenomenon associated with vascularization. The increased stiffness of colon areas may represent a new predictive marker of desmoplastic region predisposing to invasion, thus offering new potential application for monitoring adenoma with invasive potential. PMID:26940881

  10. BIOMECHANICAL and BIOCHEMICAL REMODELING of STROMAL EXTRACELLULAR MATRIX IN CANCER

    PubMed Central

    Malik, Ruchi; Lelkes, Peter I; Cukierman, Edna

    2015-01-01

    The extracellular matrix (ECM) provides structural and biochemical signals that regulate cell function. A well-controlled balance between cells and surroundings (i.e., Dynamic Reciprocity) is crucial for regulating ECM architecture. During cancer progression, epithelial cells undergo genetic alterations, which together with stromal changes, including ECM remodeling, disturb the homeostatic dynamics of the epithelium. A parallel organization of stromal ECM fibrils is associated with tumorigenic responses. In an emerging paradigm, continuous and progressive regulation via mechanical forces and aberrant signaling are believed to be responsible for tumor-associated ECM remodeling. In this review, we discuss the discrete biomechanical and biochemical mechanisms that underlie these architectural changes and highlight their particular relevance to the regulation of the alignment of ECM in the mesenchymal stroma. PMID:25708906

  11. Extracellular matrix synthesis in vascular disease: hypertension, and atherosclerosis

    PubMed Central

    Ponticos, Markella; Smith, Barbara D.

    2014-01-01

    Extracellular matrix (ECM) within the vascular network provides both a structural and regulatory role. The ECM is a dynamic composite of multiple proteins that form structures connecting cells within the network. Blood vessels are distended by blood pressure and, therefore, require ECM components with elasticity yet with enough tensile strength to resist rupture. The ECM is involved in conducting mechanical signals to cells. Most importantly, ECM regulates cellular function through chemical signaling by controlling activation and bioavailability of the growth factors. Cells respond to ECM by remodeling their microenvironment which becomes dysregulated in vascular diseases such hypertension, restenosis and atherosclerosis. This review examines the cellular and ECM components of vessels, with specific emphasis on the regulation of collagen type I and implications in vascular disease. PMID:24474961

  12. Bioactive extracellular matrix fragments in lung health and disease.

    PubMed

    Gaggar, Amit; Weathington, Nathaniel

    2016-09-01

    The extracellular matrix (ECM) is the noncellular component critical in the maintenance of organ structure and the regulation of tissue development, organ structure, and cellular signaling. The ECM is a dynamic entity that undergoes continuous degradation and resynthesis. In addition to compromising structure, degradation of the ECM can liberate bioactive fragments that cause cellular activation and chemotaxis of a variety of cells. These fragments are termed matrikines, and their cellular activities are sentinel in the development and progression of tissue injury seen in chronic lung disease. Here, we discuss the matrikines that are known to be active in lung biology and their roles in lung disease. We also consider the use of matrikines as disease markers and potential therapeutic targets in lung disease. PMID:27584731

  13. Extracellular matrix bioscaffolds in tissue remodeling and morphogenesis.

    PubMed

    Swinehart, Ilea T; Badylak, Stephen F

    2016-03-01

    During normal morphogenesis the extracellular matrix (ECM) influences cell motility, proliferation, apoptosis, and differentiation. Tissue engineers have attempted to harness the cell signaling potential of ECM to promote the functional reconstruction, if not regeneration, of injured or missing adult tissues that otherwise heal by the formation of scar tissue. ECM bioscaffolds, derived from decellularized tissues, have been used to promote the formation of site appropriate, functional tissues in many clinical applications including skeletal muscle, fibrocartilage, lower urinary tract, and esophageal reconstruction, among others. These scaffolds function by the release or exposure of growth factors and cryptic peptides, modulation of the immune response, and recruitment of progenitor cells. Herein, we describe this process of ECM induced constructive remodeling and examine similarities to normal tissue morphogenesis. PMID:26699796

  14. Cardiac extracellular matrix proteomics: Challenges, techniques, and clinical implications.

    PubMed

    Chang, Chia Wei; Dalgliesh, Ailsa J; López, Javier E; Griffiths, Leigh G

    2016-01-01

    Extracellular matrix (ECM) has emerged as a dynamic tissue component, providing not only structural support, but also functionally participating in a wide range of signaling events during development, injury, and disease remodeling. Investigation of dynamic changes in cardiac ECM proteome is challenging due to the relative insolubility of ECM proteins, which results from their macromolecular nature, extensive post-translational modification (PTM), and tendency to form protein complexes. Finally, the relative abundance of cellular and mitochondrial proteins in cardiac tissue further complicates cardiac ECM proteomic approaches. Recent developments of various techniques to enrich and analyze ECM proteins are playing a major role in overcoming these challenges. Application of cardiac ECM proteomics in disease tissues can further provide spatial and temporal information relevant to disease diagnosis, prognosis, treatment, and engineering of therapeutic candidates for cardiac repair and regeneration. PMID:26200932

  15. Characteristic adaptations of the extracellular matrix in dilated cardiomyopathy.

    PubMed

    Louzao-Martinez, Laura; Vink, Aryan; Harakalova, Magdalena; Asselbergs, Folkert W; Verhaar, Marianne C; Cheng, Caroline

    2016-10-01

    Dilated cardiomyopathy (DCM) is a relatively common heart muscle disease characterized by the dilation and thinning of the left ventricle accompanied with left ventricular systolic dysfunction. Myocardial fibrosis is a major feature in DCM and therefore it is inevitable that corresponding extracellular matrix (ECM) changes are involved in DCM onset and progression. Increasing our understanding of how ECM adaptations are involved in DCM could be important for the development of future interventions. This review article discusses the molecular adaptations in ECM composition and structure that have been reported in both animal and human studies of DCM. Furthermore, we provide a transcriptome-based catalogue of ECM genes that are associated with DCM, generated by using NCBI Gene Expression Omnibus database sets for DCM. Based on this in silico analysis, many novel ECM components involved in DCM are identified and discussed in this review. With the information gathered, we propose putative pathways of ECM adaptations in onset and progression of DCM. PMID:27391006

  16. Hypoxia and the extracellular matrix: drivers of tumour metastasis

    PubMed Central

    Gilkes, Daniele M.; Semenza, Gregg L.; Wirtz, Denis

    2014-01-01

    Of the deaths attributed to cancer, 90% are due to metastasis, and treatments that prevent or cure metastasis remain elusive. Emerging data indicate that hypoxia and the extracellular matrix (ECM) might have crucial roles in metastasis. During tumour evolution, changes in the composition and the overall content of the ECM reflect both its biophysical and biological properties and these strongly influence tumour and stromal cell properties, such as proliferation and motility. Originally thought of as independent contributors to metastatic spread, recent studies have established a direct link between hypoxia and the composition and the organization of the ECM, which suggests a new model in which multiple microenvironmental signals might converge to synergistically influence metastatic outcome. PMID:24827502

  17. ADAMTS: a novel family of extracellular matrix proteases.

    PubMed

    Tang, B L

    2001-01-01

    ADAMTS (a disintegrin and metalloprotease with thrombospondin motifs) is a novel family of extracellular proteases found in both mammals and invertebrates. Members of the family may be distinguished from the ADAM (a disintegrin and metalloprotease) family members based on the multiple copies of thrombospondin 1-like repeats they carry. With at least nine members in mammals alone, the ADAMTS family members are predicted by their structural domains to be extracellular matrix (ECM) proteins with a wide range of activities and functions distinct from members of the ADAM family that are largely anchored on the cell surface. ADAMTS2 is a procollagen N-proteinase, and the mutations of its gene are responsible for Human Ehlers-Danlos syndrome type VII C and bovine dermatosparaxis. ADAMTS4 and ADAMTS5 are aggrecanases implicated in the degradation of cartilage aggrecan in arthritic diseases. Other members of the ADAMTS family have also been implicated in roles during embryonic development and angiogenesis. Current and future studies on this emerging group of ECM proteases may provide important insights into developmental or pathological processes involving ECM remodeling. PMID:11167130

  18. Cells involved in extracellular matrix remodeling after acute myocardial infarction

    PubMed Central

    Garcia, Larissa Ferraz; Mataveli, Fábio D’Aguiar; Mader, Ana Maria Amaral Antônio; Theodoro, Thérèse Rachell; Justo, Giselle Zenker; Pinhal, Maria Aparecida da Silva

    2015-01-01

    Objective Evaluate the effects of VEGF165 gene transfer in the process of remodeling of the extracellular matrix after an acute myocardial infarct. Methods Wistar rats were submitted to myocardial infarction, after the ligation of the left descending artery, and the left ventricle ejection fraction was used to classify the infarcts into large and small. The animals were divided into groups of ten, according to the size of infarcted area (large or small), and received or not VEGF165 treatment. Evaluation of different markers was performed using immunohistochemistry and digital quantification. The primary antibodies used in the analysis were anti-fibronectin, anti-vimentin, anti-CD44, anti-E-cadherin, anti-CD24, anti-alpha-1-actin, and anti-PCNA. The results were expressed as mean and standard error, and analyzed by ANOVA, considering statistically significant if p≤0.05. Results There was a significant increase in the expression of undifferentiated cell markers, such as fibronectin (protein present in the extracellular matrix) and CD44 (glycoprotein present in the endothelial cells). However, there was decreased expression of vimentin and PCNA, indicating a possible decrease in the process of cell proliferation after treatment with VEGF165. Markers of differentiated cells, E-cadherin (adhesion protein between myocardial cells), CD24 (protein present in the blood vessels), and alpha-1-actin (specific myocyte marker), showed higher expression in the groups submitted to gene therapy, compared to non-treated group. The value obtained by the relation between alpha-1-actin and vimentin was approximately three times higher in the groups treated with VEGF165, suggesting greater tissue differentiation. Conclusion The results demonstrated the important role of myocytes in the process of tissue remodeling, confirming that VEGF165 seems to provide a protective effect in the treatment of acute myocardial infarct. PMID:25993074

  19. Binding of the extracellular matrix component entactin to Candida albicans.

    PubMed Central

    López-Ribot, J L; Chaffin, W L

    1994-01-01

    We have investigated the interaction between Candida albicans and entactin, a recently characterized glycoprotein present in the extracellular matrix, especially in the basement membrane. Organisms of both the yeast and the hyphal morphologies of the fungus had the ability to bind recombinant entactin, as detected by an indirect immunofluorescence assay. Material present in the 2-mercaptoethanol cell wall extracts from both C. albicans growth forms was capable of binding to immobilized recombinant entactin in a dose-dependent manner. Binding to entactin was approximately twice that observed for laminin. Binding of an extract component(s) to entactin was partially inhibited by an Arg-Gly-Asp-Ser peptide. A polyclonal antientactin antiserum, as well as a pooled antiserum preparation raised against components present in different C. albicans cell wall extracts, completely or almost completely abolished binding. The existence of morphology-specific receptor-like molecules which bind to different domains of the entactin molecule was ruled out in a competition binding assay. The entactin-binding material(s) in the cell wall also displayed some ability to bind laminin and fibronectin, since preadsorption in the presence of these extracellular matrix components resulted in reduction of binding to entactin. Moieties with a molecular mass of approximately 25, 44, and 65 kDa present in the 2-mercaptoethanol cell wall extracts from both blastoconidia and germ tubes were detected in a ligand affinity blotting experiment as having the ability to bind entactin. Interactions between C. albicans and entactin could be important in mediating adhesion of the fungus to the host tissues and may play a role in the establishment of the disseminated form of the disease. Images PMID:7927722

  20. SPARC mediates early extracellular matrix remodeling following myocardial infarction

    PubMed Central

    McCurdy, Sarah M.; Dai, Qiuxia; Zhang, Jianhua; Zamilpa, Rogelio; Ramirez, Trevi A.; Dayah, Tariq; Nguyen, Nguyen; Jin, Yu-Fang; Bradshaw, Amy D.

    2011-01-01

    Secreted protein, acidic, and rich in cysteine (SPARC) is a matricellular protein that functions in the extracellular processing of newly synthesized collagen. Collagen deposition to form a scar is a key event following a myocardial infarction (MI). Because the roles of SPARC in the early post-MI setting have not been defined, we examined age-matched wild-type (WT; n=22) and SPARC-deficient (null; n=25) mice at day 3 post-MI. Day 0 WT (n=28) and null (n=20) mice served as controls. Infarct size was 52 ± 2% for WT and 47 ± 2% for SPARC null (P=NS), indicating that the MI injury was comparable in the two groups. By echocardiography, WT mice increased end-diastolic volumes from 45 ± 2 to 83 ± 5 μl (P < 0.05). SPARC null mice also increased end-diastolic volumes but to a lesser extent than WT (39 ± 3 to 63 ± 5 μl; P < 0.05 vs. day 0 controls and vs. WT day 3 MI). Ejection fraction fell post-MI in WT mice from 57 ± 2 to 19 ± 1%. The decrease in ejection fraction was attenuated in the absence of SPARC (65 ± 2 to 28 ± 2%). Fibroblasts isolated from SPARC null left ventricle (LV) showed differences in the expression of 22 genes encoding extracellular matrix and adhesion molecule genes, including fibronectin, connective tissue growth factor (CTGF; CCN2), matrix metalloproteinase-3 (MMP-3), and tissue inhibitor of metalloproteinase-2 (TIMP-2). The change in fibroblast gene expression levels was mirrored in tissue protein extracts for fibronectin, CTGF, and MMP-3 but not TIMP-2. Combined, the results of this study indicate that SPARC deletion preserves LV function at day 3 post-MI but may be detrimental for the long-term response due to impaired fibroblast activation. PMID:21602472

  1. Hydrogels derived from demineralized and decellularized bone extracellular matrix

    PubMed Central

    Sawkins, M.J.; Bowen, W.; Dhadda, P.; Markides, H.; Sidney, L.E.; Taylor, A.J.; Rose, F.R.A.J.; Badylak, S.F.; Shakesheff, K.M.; White, L.J.

    2013-01-01

    The extracellular matrix (ECM) of mammalian tissues has been isolated, decellularized and utilized as a scaffold to facilitate the repair and reconstruction of numerous tissues. Recent studies have suggested that superior function and complex tissue formation occurred when ECM scaffolds were derived from site-specific homologous tissues compared with heterologous tissues. The objectives of the present study were to apply a stringent decellularization process to demineralized bone matrix (DBM), prepared from bovine bone, and to characterize the structure and composition of the resulting ECM materials and DBM itself. Additionally, we sought to produce a soluble form of DBM and ECM which could be induced to form a hydrogel. Current clinical delivery of DBM particles for treatment of bone defects requires incorporation of the particles within a carrier liquid. Differences in osteogenic activity, inflammation and nephrotoxicity have been reported with various carrier liquids. The use of hydrogel forms of DBM or ECM may reduce the need for carrier liquids. DBM and ECM hydrogels exhibited sigmoidal gelation kinetics consistent with a nucleation and growth mechanism, with ECM hydrogels characterized by lower storage moduli than the DBM hydrogels. Enhanced proliferation of mouse primary calvarial cells was achieved on ECM hydrogels, compared with collagen type I and DBM hydrogels. These results show that DBM and ECM hydrogels have distinct structural, mechanical and biological properties and have the potential for clinical delivery without the need for carrier liquids. PMID:23624219

  2. Extracellular matrix-mediated chemotaxis can impede cell migration

    PubMed Central

    Perumpanani, A. J.; Simmons, D. L.; Gearing, A. J. H.; Miller, K. M.; Ward, G.; Norbury, J.; Schneemann, M.; Sherratt, J. A.

    1998-01-01

    Cells use a combination of changes in adhesion, proteolysis and motility (directed and random) during the process of migration. Proteolysis of the extracellular matrix (ECM) results in thecreation of haptotactic gradients which cells use to move in a directed fashion. The proteolytic creation of these gradients also results in the production of digested fragments of ECM. In this study we show that in the human fibrosarcoma cell line HT1080, matrix metalloproteinase-2(MMP-2)-digested fragments of fibronectin exert a chemotactic pull stronger than that of undigested fibronectin. During invasion, this gradient of ECM fragments is established in the wake of an invading cell, running counter to the direction of invasion. The resultant chemotactic pull is anti-invasive, contrary to the traditional view of the role of chemotaxis in invasion. Uncontrolled ECM degradation by high concentrations of MMP can thus result in steep gradients of ECM fragments, which run against the direction of invasion. Consequently, the invasive potential of a cell depends on MMP production in a biphasic mannerimplying that MMP inhibitors will upregulate invasion in high-MMPexpressing cells. Hence the therapeutic use of protease inhibitors against tumours expressing high levels of MMP could produce an augmentation of invasion.

  3. Dynamic Interactions of Epidermal Collagen XVII with the Extracellular Matrix

    PubMed Central

    Nishie, Wataru; Kiritsi, Dimitra; Nyström, Alexander; Hofmann, Silke C.; Bruckner-Tuderman, Leena

    2011-01-01

    Transmembrane collagen XVII, a major component of the hemidesmosomes, is crucial for stable adhesion of the epidermis and dermis in the skin, and its dysfunction results in blistering diseases. The ectodomain of collagen XVII (Ecto-ColXVII) is constitutively shed from the cell surface, but its binding partner(s) in the extracellular matrix (ECM) and the physiologic roles of the ligand interactions remain elusive. Herein, we used a new cleavage site–specific antibody to address the dynamics of collagen XVII shedding and the interactions of Ecto-ColXVII with the ECM. Ecto-ColXVII was present in the migration tracks of primary human keratinocytes and co-localized with laminin 332. The presence of this laminin, but also of collagen IV and Matrigel, in the ECM enhanced shedding and incorporation of Ecto-ColXVII into the matrix. Laminin 332 is a major, but not exclusive, interaction partner in vivo because Ecto-ColXVII deposited in the ECM of laminin 332–deficient keratinocytes was drastically reduced, but Ecto-ColXVII was present in laminin 332–negative human skin. Expression of collagen XVII deletion mutants in HEK 293 cells identified the C-terminal ectodomain stretch Ser978-Pro1497 as necessary for ECM binding. Taken together, migrating keratinocytes shed the Ecto-ColXVII, and this dynamically binds via its C-terminal domain to distinct partners in the ECM. PMID:21801871

  4. The cytoprotective capacity of processed human cardiac extracellular matrix.

    PubMed

    Kappler, Benjamin; Anic, Petra; Becker, Matthias; Bader, Andreas; Klose, Kristin; Klein, Oliver; Oberwallner, Barbara; Choi, Yeong-Hoon; Falk, Volkmar; Stamm, Christof

    2016-07-01

    Freshly isolated human cardiac extracellular matrix sheets (cECM) have been shown to support stem cell proliferation and tissue-specific lineage commitment. We now developed a protocol for standardized production of durable, bio-functional hcECM microparticles and corresponding hydrogel, and tested its cytoprotective effects on contractile cells subjected to ischemia-like conditions. Human ventricular myocardium was decellularized by a 3-step protocol, including Tris/EDTA, SDS and serum incubation (cECM). Following snap-freezing and lyophilization, microparticles were created and characterized by laser diffraction, dynamic image analysis (DIA), and mass spectrometry. Moreover, cECM hydrogel was produced by pepsin digestion. Baseline cell-support characteristics were determined using murine HL-1 cardiomyocytes, and the cytoprotective effects of ECM products were tested under hypoxia and glucose/serum deprivation. In cECM, glycoproteins (thrombospondin 1, fibronectin, collagens and nidogen-1) and proteoglycans (dermatopontin, lumican and mimecan) were preserved, but residual intracellular and blood-borne proteins were also detected. The median particle feret diameter was 66 μm (15-157 μm) by laser diffraction, and 57 μm (20-182 μm) by DIA with crystal violet staining. HL-1 cells displayed enhanced metabolic activity (39 ± 12 %, P < 0.05) and proliferation (16 ± 3 %, P < 0.05) when grown on cECM microparticles in normoxia. During simulated ischemia, cECM microparticles exerted distinct cytoprotective effects (MTS conversion, 240 ± 32 %; BrdU uptake, 45 ± 14 %; LDH release, -72 ± 7 %; P < 0.01, each). When cECM microparticles were solubilized to form a hydrogel, the cytoprotective effect was initially abolished. However, modifying the preparation process (pepsin digestion at pH 2 and 25 °C, 1 mg/ml final cECM concentration) restored the cytoprotective cECM activity. Extracellular matrix from human myocardium can be processed to

  5. The tetrapartite synapse: Extracellular matrix remodeling contributes to corticoaccumbens plasticity underlying drug addiction.

    PubMed

    Smith, Alexander C W; Scofield, Michael D; Kalivas, Peter W

    2015-12-01

    Synaptic plasticity has long been known to involve three key elements of neuropil, the presynapse, the postsynapse and adjacent glia. Here we review the role of the extracellular matrix in synaptic plasticity as a necessary component forming the tetrapartite synapse. We describe the role of matrix metalloproteinases as enzymes sculpting extracellular proteins and thereby creating an extracellular signaling domain required for synaptic plasticity. Specifically we focus on the role of the tetrapartite synapse in mediating the effects of addictive drugs at cortico-striatal synapses, and conclude that the extracellular signaling domain and its regulation by matrix metalloproteinases is critical for developing and expressing drug seeking behaviors. PMID:25838241

  6. Small intestinal submucosa extracellular matrix (CorMatrix®) in cardiovascular surgery: a systematic review.

    PubMed

    Mosala Nezhad, Zahra; Poncelet, Alain; de Kerchove, Laurent; Gianello, Pierre; Fervaille, Caroline; El Khoury, Gebrine

    2016-06-01

    Extracellular matrix (ECM) derived from small intestinal submucosa (SIS) is widely used in clinical applications as a scaffold for tissue repair. Recently, CorMatrix® porcine SIS-ECM (CorMatrix Cardiovascular, Inc., Roswell, GA, USA) has gained popularity for 'next-generation' cardiovascular tissue engineering due to its ease of use, remodelling properties, lack of immunogenicity, absorbability and potential to promote native tissue growth. Here, we provide an overview of the biology of porcine SIS-ECM and systematically review the preclinical and clinical literature on its use in cardiovascular surgery. CorMatrix® has been used in a variety of cardiovascular surgical applications, and since it is the most widely used SIS-ECM, this material is the focus of this review. Since CorMatrix® is a relatively new product for cardiovascular surgery, some clinical and preclinical studies published lack systematic reporting of functional and pathological findings in sufficient numbers of subjects. There are also emerging reports to suggest that, contrary to expectations, an undesirable inflammatory response may occur in CorMatrix® implants in humans and longer-term outcomes at particular sites, such as the heart valves, may be suboptimal. Large-scale clinical studies are needed driven by robust protocols that aim to quantify the pathological process of tissue repair. PMID:26912574

  7. Extracellular matrix turnover in coronary artery ectasia patients.

    PubMed

    Liu, Ruifeng; Chen, Lianfeng; Wu, Wei; Chen, Houzao; Zhang, Shuyang

    2016-03-01

    Dysregulation of the metabolism of the extracellular matrix (ECM) may contribute to coronary artery ectasia (CAE). This study evaluated the turnover of main ECM components and related proteolytic enzymes activities. In this study, thirty patients with CAE, 30 patients with coronary artery disease (CAD) and 30 subjects with normal coronary arteries (Control) were selected. The following circulating ECM metabolism markers were measured: soluble elastin (sElastin), collagen type I cross-linked telopeptides (ICTP), procollagen type I carboxy terminal peptide (PICP), protocollagen III N-terminal propeptide (PIIINP), and procollagen a1(III) C-terminal propeptide (PIIICP). Serum total elastase activity and total matrix metalloproteinase (MMP) activity were also determined. The level of sElastin was higher in the CAE group than in the CAD and Control groups (P1 = 0.009, P2 = 0.000). There was no difference in ICTP (P = 0.168) or PIIICP (P = 0.079) among the three groups. PICP was significantly elevated in CAE (P1 = 0.001, P2 = 0.002). PIIINP was also significantly increased in CAE (P1 = 0.002, P2 = 0.007). Total elastase activity was higher in the CAE group than in the other two groups (P1 = 0.006, P2 = 0.022). Total MMP activity was significantly higher in the CAE group than the Control group (P2 = 0.013) but not higher than the CAD group (P1 = 0.477). In conclusion, within CAE patients the main changes in ECM metabolism were increased degradation of elastin fibres and the transition of collagen from type III to type I. Elastase and MMPs appear to be associated with this kind of ECM turnover. PMID:25576491

  8. Astrocytes as a Source for Extracellular Matrix Molecules and Cytokines

    PubMed Central

    Wiese, Stefan; Karus, Michael; Faissner, Andreas

    2012-01-01

    Research of the past 25 years has shown that astrocytes do more than participating and building up the blood-brain barrier and detoxify the active synapse by reuptake of neurotransmitters and ions. Indeed, astrocytes express neurotransmitter receptors and, as a consequence, respond to stimuli. Within the tripartite synapse, the astrocytes owe more and more importance. Besides the functional aspects the differentiation of astrocytes has gained a more intensive focus. Deeper knowledge of the differentiation processes during development of the central nervous system might help explaining and even help treating neurological diseases like Alzheimer’s disease, Amyotrophic lateral sclerosis, Parkinsons disease, and psychiatric disorders in which astrocytes have been shown to play a role. Specific differentiation of neural stem cells toward the astroglial lineage is performed as a multi-step process. Astrocytes and oligodendrocytes develop from a multipotent stem cell that prior to this has produced primarily neuronal precursor cells. This switch toward the more astroglial differentiation is regulated by a change in receptor composition on the cell surface and responsiveness to Fibroblast growth factor and Epidermal growth factor (EGF). The glial precursor cell is driven into the astroglial direction by signaling molecules like Ciliary neurotrophic factor, Bone Morphogenetic Proteins, and EGF. However, the early astrocytes influence their environment not only by releasing and responding to diverse soluble factors but also express a wide range of extracellular matrix (ECM) molecules, in particular proteoglycans of the lectican family and tenascins. Lately these ECM molecules have been shown to participate in glial development. In this regard, especially the matrix protein Tenascin C (Tnc) proved to be an important regulator of astrocyte precursor cell proliferation and migration during spinal cord development. Nevertheless, ECM molecules expressed by reactive astrocytes

  9. Extracellular matrix protein expression is brain region dependent.

    PubMed

    Dauth, Stephanie; Grevesse, Thomas; Pantazopoulos, Harry; Campbell, Patrick H; Maoz, Ben M; Berretta, Sabina; Parker, Kevin Kit

    2016-05-01

    In the brain, extracellular matrix (ECM) components form networks that contribute to structural and functional diversity. Maladaptive remodeling of ECM networks has been reported in neurodegenerative and psychiatric disorders, suggesting that the brain microenvironment is a dynamic structure. A lack of quantitative information about ECM distribution in the brain hinders an understanding of region-specific ECM functions and the role of ECM in health and disease. We hypothesized that each ECM protein as well as specific ECM structures, such as perineuronal nets (PNNs) and interstitial matrix, are differentially distributed throughout the brain, contributing to the unique structure and function in the various regions of the brain. To test our hypothesis, we quantitatively analyzed the distribution, colocalization, and protein expression of aggrecan, brevican, and tenascin-R throughout the rat brain utilizing immunohistochemistry and mass spectrometry analysis and assessed the effect of aggrecan, brevican, and/or tenascin-R on neurite outgrowth in vitro. We focused on aggrecan, brevican, and tenascin-R as they are especially expressed in the mature brain, and have established roles in brain development, plasticity, and neurite outgrowth. The results revealed a differentiated distribution of all three proteins throughout the brain and indicated that their presence significantly reduces neurite outgrowth in a 3D in vitro environment. These results underline the importance of a unique and complex ECM distribution for brain physiology and suggest that encoding the distribution of distinct ECM proteins throughout the brain will aid in understanding their function in physiology and in turn assist in identifying their role in disease. J. Comp. Neurol. 524:1309-1336, 2016. © 2016 Wiley Periodicals, Inc. PMID:26780384

  10. Extracellular matrix stiffness dictates Wnt expression through integrin pathway

    PubMed Central

    Du, Jing; Zu, Yan; Li, Jing; Du, Shuyuan; Xu, Yipu; Zhang, Lang; Jiang, Li; Wang, Zhao; Chien, Shu; Yang, Chun

    2016-01-01

    It is well established that extracellular matrix (ECM) stiffness plays a significant role in regulating the phenotypes and behaviors of many cell types. However, the mechanism underlying the sensing of mechanical cues and subsequent elasticity-triggered pathways remains largely unknown. We observed that stiff ECM significantly enhanced the expression level of several members of the Wnt/β-catenin pathway in both bone marrow mesenchymal stem cells and primary chondrocytes. The activation of β-catenin by stiff ECM is not dependent on Wnt signals but is elevated by the activation of integrin/ focal adhesion kinase (FAK) pathway. The accumulated β-catenin then bound to the wnt1 promoter region to up-regulate the gene transcription, thus constituting a positive feedback of the Wnt/β-catenin pathway. With the amplifying effect of positive feedback, this integrin-activated β-catenin/Wnt pathway plays significant roles in mediating the enhancement of Wnt signal on stiff ECM and contributes to the regulation of mesenchymal stem cell differentiation and primary chondrocyte phenotype maintenance. The present integrin-regulated Wnt1 expression and signaling contributes to the understanding of the molecular mechanisms underlying the regulation of cell behaviors by ECM elasticity. PMID:26854061

  11. EVALUATION OF THE EXTRACELLULAR MATRIX OF INJURED SUPRASPINATUS IN RATS

    PubMed Central

    Almeida, Luiz Henrique Oliveira; Ikemoto, Roberto; Mader, Ana Maria; Pinhal, Maria Aparecida Silva; Munhoz, Bruna; Murachovsky, Joel

    2016-01-01

    ABSTRACT Objective: To evaluate the evolution of injuries of the supraspinatus muscle by immunohistochemistry (IHC) and anatomopathological analysis in animal model (Wistar rats). Methods: Twenty-five Wistar rats were submitted to complete injury of the supraspinatus tendon, then subsequently sacrificed in groups of five animals at the following periods: immediately after the injury, 24h after the injury, 48h after, 30 days after and three months after the injury. All groups underwent histological and IHC analysis. Results: Regarding vascular proliferation and inflammatory infiltrate, we found a statistically significant difference between groups 1(control group) and 2 (24h after injury). IHC analysis showed that expression of vascular endothelial growth factor (VEGF) showed a statistically significant difference between groups 1 and 2, and collagen type 1 (Col-1) evaluation presented a statistically significant difference between groups 1 and 4. Conclusion: We observed changes in the extracellular matrix components compatible with remodeling and healing. Remodeling is more intense 24h after injury. However, VEGF and Col-1 are substantially increased at 24h and 30 days after the injury, respectively. Level of Evidence I, Experimental Study. PMID:26997907

  12. Extracellular matrix, biotensegrity and tumor microenvironment. An update and overview.

    PubMed

    Noguera, R; Nieto, O A; Tadeo, I; Fariñas, F; Alvaro, T

    2012-06-01

    The extracellular matrix (ECM) constitutes a three-dimensional network that surrounds all cells, organs and tissues in the body. It forms a biophysical filter for protection, nutrition and cell innervation, as well as the medium for facilitating immune response, angiogenesis, fibrosis and tissue regeneration. It is the mechanism by which mechanical forces are transmitted to the basement membrane which, through the integrins, supports the tensegrity system and activates the epigenetic mechanisms of the cell. A review and update on current knowledge on this topic reveals how disturbance of the ECM leads to a loss of efficient filtering, nutrition, elimination, and cell denervation functions, in addition to loss of regeneration capacity and disorders in mechanotransduction. Furthermore, such disturbance results in a loss of substrate, and with it the ability to provide a proper immune response against tumor, toxic and infectious agents. Reciprocal communication between ECM stromal and parenchymatous cells directs gene expression. The oncogenic capacity of the stroma derives from the associated cells as well as from the tumor cells, the angiogenic microenvironment and from an alteration in tensegrity; all of which are dependent on the ECM. It has been shown that the malignant phenotype is reversible by correction of the altered cues of the ECM. PMID:22473691

  13. Tissue Extracellular Matrix Nanoparticle Presentation in Electrospun Nanofibers

    PubMed Central

    Gibson, Matt; Mao, Hai-Quan; Elisseeff, Jennifer

    2014-01-01

    Biomaterials derived from the decellularization of mature tissues retain biological and architectural features that profoundly influence cellular activity. However, the clinical utility of such materials remains limited as the shape and physical properties are difficult to control. In contrast, scaffolds based on synthetic polymers can be engineered to exhibit specific physical properties, yet often suffer from limited biological functionality. This study characterizes composite materials that present decellularized extracellular matrix (DECM) particles in combination with synthetic nanofibers and examines the ability of these materials to influence stem cell differentiation. Mechanical processing of decellularized tissues yielded particles with diameters ranging from 71 to 334 nm. Nanofiber scaffolds containing up to 10% DECM particles (wt/wt) derived from six different tissues were engineered and evaluated to confirm DECM particle incorporation and to measure bioactivity. Scaffolds containing bone, cartilage, and fat promoted osteogenesis at 1 and 3 weeks compared to controls. In contrast, spleen and lung DECM significantly reduced osteogenic outcomes compared to controls. These findings highlight the potential to incorporate appropriate source DECM nanoparticles within nanofiber composites to design a scaffold with bioactivity targeted to specific applications. PMID:24971329

  14. Fibroblast extracellular matrix and adhesion on microtextured polydimethylsiloxane scaffolds.

    PubMed

    Stanton, Morgan M; Parrillo, Allegra; Thomas, Gawain M; McGimpsey, W Grant; Wen, Qi; Bellin, Robert M; Lambert, Christopher R

    2015-05-01

    The immediate physical and chemical surroundings of cells provide important biochemical cues for their behavior. Designing and tailoring biomaterials for controlled cell signaling and extracellular matrix (ECM) can be difficult due to the complexity of the cell-surface relationship. To address this issue, our research has led to the development of a polydimethylsiloxane (PDMS) scaffold with defined microtopography and chemistry for surface driven ECM assembly. When human fibroblasts were cultured on this microtextured PDMS with 2-6 µm wide vertical features, significant changes in morphology, adhesion, actin cytoskeleton, and fibronectin generation were noted when compared with cells cultured on unmodified PDMS. Investigation of cellular response and behavior was performed with atomic force microscopy in conjunction with fluorescent labeling of focal adhesion cites and fibronectin in the ECM. Changes in the surface topography induced lower adhesion, an altered actin cytoskeleton, and compacted units of fibronectin similar to that observed in vivo. Overall, these findings provide critical information of cell-surface interactions with a microtextured, polymer substrate that can be used in the field of tissue engineering for controlling cellular ECM interactions. PMID:25142015

  15. Cellular and extracellular matrix modulation of corneal stromal opacity.

    PubMed

    Torricelli, Andre A M; Wilson, Steven E

    2014-12-01

    Stromal transparency is a critical factor contributing to normal function of the visual system. Corneal injury, surgery, disease and infection elicit complex wound healing responses that serve to protect against insults and maintain the integrity of the cornea, and subsequently to restore corneal structure and transparency. However, in some cases these processes result in prolonged loss of corneal transparency and resulting diminished vision. Corneal opacity is mediated by the complex actions of many cytokines, growth factors, and chemokines produced by the epithelial cells, stromal cells, bone marrow-derived cells, lacrimal tissues, and nerves. Myofibroblasts, and the disorganized extracellular matrix produced by these cells, are critical determinants of the level and persistence of stromal opacity after corneal injury. Decreases in corneal crystallins in myofibroblasts and corneal fibroblasts contribute to cellular opacity in the stroma. Regeneration of a fully functional epithelial basement membrane (BM) appears to have a critical role in the maintenance of corneal stromal transparency after mild injuries and recovery of transparency when opacity is generated after severe injuries. The epithelial BM likely has a regulatory function whereby it modulates epithelium-derived growth factors such as transforming growth factor (TGF) β and platelet-derived growth factor (PDGF) that drive the development and persistence of myofibroblasts from precursor cells. The purpose of this article is to review the factors involved in the maintenance of corneal transparency and to highlight the mechanisms involved in the appearance, persistency and regression of corneal opacity after stromal injury. PMID:25281830

  16. Extracellular matrix stiffness dictates Wnt expression through integrin pathway.

    PubMed

    Du, Jing; Zu, Yan; Li, Jing; Du, Shuyuan; Xu, Yipu; Zhang, Lang; Jiang, Li; Wang, Zhao; Chien, Shu; Yang, Chun

    2016-01-01

    It is well established that extracellular matrix (ECM) stiffness plays a significant role in regulating the phenotypes and behaviors of many cell types. However, the mechanism underlying the sensing of mechanical cues and subsequent elasticity-triggered pathways remains largely unknown. We observed that stiff ECM significantly enhanced the expression level of several members of the Wnt/β-catenin pathway in both bone marrow mesenchymal stem cells and primary chondrocytes. The activation of β-catenin by stiff ECM is not dependent on Wnt signals but is elevated by the activation of integrin/ focal adhesion kinase (FAK) pathway. The accumulated β-catenin then bound to the wnt1 promoter region to up-regulate the gene transcription, thus constituting a positive feedback of the Wnt/β-catenin pathway. With the amplifying effect of positive feedback, this integrin-activated β-catenin/Wnt pathway plays significant roles in mediating the enhancement of Wnt signal on stiff ECM and contributes to the regulation of mesenchymal stem cell differentiation and primary chondrocyte phenotype maintenance. The present integrin-regulated Wnt1 expression and signaling contributes to the understanding of the molecular mechanisms underlying the regulation of cell behaviors by ECM elasticity. PMID:26854061

  17. The significance of the extracellular matrix in intracranial aneurysms.

    PubMed

    Austin, G; Fisher, S; Dickson, D; Anderson, D; Richardson, S

    1993-01-01

    To what extent might change sin the extracellular matrix proteins in intracranial aneurysms (IA) or parent artery (PA) wall contribute to enlargement? Using IA specimens obtained from the coroner's office, the wall structure by light (Trichrome and Actin stains) and immunofluorescent microscopy (IF) (collagen I and fibronectin) have been studied in 18 specimens. It is well known that IA only arise when there is a deficiency in the internal elastic membrane and some breakdown in the media. The results show a normal three layer structure of intima, media, (muscularis), and adventitia in the PA extending out into the neck of the aneurysm. Immunofluorescent stains (IS) show collagen I and fibronectin limited to the adventitia and muscularis, respectively, of the PA and control arteries. The enlarging IA wall shows almost complete loss of this architecture. By IF stain, collagen I and fibronectin are intermingled throughout the IA wall. It is postulated that loss of structure and the confining muscularis layer permit easier elastic enlargement from a more or less homogeneous wall structure. PMID:7681275

  18. The extracellular matrix as a cell survival factor.

    PubMed Central

    Meredith, J E; Fazeli, B; Schwartz, M A

    1993-01-01

    Programmed cell death (PCD) or apoptosis is a naturally occurring cell suicide pathway induced in a variety of cell types. In many cases, PCD is induced by the withdrawal of specific hormones or growth factors that function as survival factors. In this study, we have investigated the potential role of the extracellular matrix (ECM) as a cell survival factor. Our results indicate that in the absence of any ECM interactions, human endothelial cells rapidly undergo PCD, as determined by cell morphology, nuclei fragmentation, DNA degradation, protein cross-linking, and the expression of the PCD-specific gene TRPM-2. PCD was blocked by plating cells on an immobilized integrin beta 1 antibody but not by antibodies to either the class I histocompatibility antigen (HLA) or vascular cell adhesion molecule-1 (VCAM-1), suggesting that integrin-mediated signals were required for maintaining cell viability. Treatment of the cells in suspension with the tyrosine phosphatase inhibitor sodium orthovanadate also blocked PCD. When other cell types were examined, some, but not all, underwent rapid cell death when deprived of adhesion to the ECM. These results suggest that in addition to regulating cell growth and differentiation, the ECM also functions as a survival factor for many cell types. Images PMID:8257797

  19. Expression of genes encoding extracellular matrix proteins: A macroarray study

    PubMed Central

    FUTYMA, KONRAD; MIOTŁA, PAWEŁ; RÓŻYŃSKA, KRYSTYNA; ZDUNEK, MAŁGORZATA; SEMCZUK, ANDRZEJ; RECHBERGER, TOMASZ; WOJCIEROWSKI, JACEK

    2014-01-01

    Endometrial cancer (EC) is one of the most common gynecological malignancies in Poland, with well-established risk factors. Genetic instability and molecular alterations responsible for endometrial carcinogenesis have been systematically investigated. The aim of the present study was to investigate, by means of cDNA macroarrays, the expression profiles of genes encoding extracellular matrix (ECM) proteins in ECs. Tissue specimens were collected during surgical procedures from 40 patients with EC, and control tissue was collected from 9 patients with uterine leiomyomas. RNA was isolated and RT-PCR with radioisotope-labeled cDNA was performed. The levels of ECM protein gene expression in normal endometrial tissues were compared to the expression of these genes in EC specimens. Statistically significant differences in gene expression, stratified by clinical stage of the ECs, were detected for aggrecan, vitronectin, tenascin R, nidogen and two collagen proteins: type VIII chain α1 and type XI chain α2. All of these proteins were overexpressed in stage III endometrial carcinomas compared to levels in stage I and II uterine neoplasms. In conclusion, increased expression of genes encoding ECM proteins may play an important role in facilitating accelerated disease progression of human ECs. PMID:25231141

  20. Using Extracellular Matrix Proteomics: To Understand Left Ventricular Remodeling

    PubMed Central

    Lindsey, Merry L.; Weintraub, Susan T.; Lange, Richard A.

    2011-01-01

    Survival following myocardial infarction (MI) has improved substantially over the last 40 years; however, the incidence of subsequent congestive heart failure has dramatically increased as a consequence. Discovering plasma markers that signify adverse cardiac remodeling may allow high-risk patients to be recognized earlier and may provide an improved way to assess treatment efficacy. Alterations in extracellular matrix (ECM) regulate cardiac remodeling following MI and potentially provide a large array of candidate indicators. The field of cardiac proteomics has progressed rapidly over the past 20 years, since publication of the first two-dimensional electrophoretic gels of left ventricle proteins. Proteomic approaches are now routinely utilized to better understand how the left ventricle responds to injury. In this review, we will discuss how methods have developed to allow comprehensive evaluation of the ECM proteome. We will explain how ECM proteomic data can be used to predict adverse remodeling for an individual patient and highlight future directions. Although this review will focus on the use of ECM proteomics to better understand post-MI remodeling responses, these approaches have applicability to a wide-range of cardiac pathologies, including pressure overload hypertrophy, viral myocarditis, and non-ischemic heart failure. PMID:22337931

  1. Substrate stiffness regulates extracellular matrix deposition by alveolar epithelial cells

    PubMed Central

    Eisenberg, Jessica L; Safi, Asmahan; Wei, Xiaoding; Espinosa, Horacio D; Budinger, GR Scott; Takawira, Desire; Hopkinson, Susan B; Jones, Jonathan CR

    2012-01-01

    Aim The aim of the study was to address whether a stiff substrate, a model for pulmonary fibrosis, is responsible for inducing changes in the phenotype of alveolar epithelial cells (AEC) in the lung, including their deposition and organization of extracellular matrix (ECM) proteins. Methods Freshly isolated lung AEC from male Sprague Dawley rats were seeded onto polyacrylamide gel substrates of varying stiffness and analyzed for expression and organization of adhesion, cytoskeletal, differentiation, and ECM components by Western immunoblotting and confocal immunofluorescence microscopy. Results We observed that substrate stiffness influences cell morphology and the organization of focal adhesions and the actin cytoskeleton. Surprisingly, however, we found that substrate stiffness has no influence on the differentiation of type II into type I AEC, nor does increased substrate stiffness lead to an epithelial–mesenchymal transition. In contrast, our data indicate that substrate stiffness regulates the expression of the α3 laminin subunit by AEC and the organization of both fibronectin and laminin in their ECM. Conclusions An increase in substrate stiffness leads to enhanced laminin and fibronectin assembly into fibrils, which likely contributes to the disease phenotype in the fibrotic lung. PMID:23204878

  2. Mechanocompatible Polymer-Extracellular-Matrix Composites for Vascular Tissue Engineering.

    PubMed

    Jiang, Bin; Suen, Rachel; Wang, Jiao-Jing; Zhang, Zheng J; Wertheim, Jason A; Ameer, Guillermo A

    2016-07-01

    Small-diameter vascular grafts developed from vascular extracellular matrix (ECM) can potentially be used for bypass surgeries and other vascular reconstruction and repair procedures. The addition of heparin to the ECM improves graft hemocompatibility but often involves chemical cross-linking, which increases ECM mechanical stiffness compared to native arteries. Herein, the importance of maintaining ECM mechanocompatibility is demonstrated, and a mechanocompatible strategy to immobilize heparin onto the ECM via a biodegradable elastomer is described. Specifically, poly(1,8-octamethylene citrate)-co-cysteine is hybridized to the ECM, forming a polymer-ECM composite that allows for heparin immobilization via maleimide-thiol "click" chemistry. Heparinized composites reduce platelet adhesion by >60% in vitro, without altering the elastic modulus of the ECM. In a rat abdominal aortic interposition model, intimal hyperplasia in heparinized mechanocompatible grafts is 65% lower when compared to ECM-only control grafts at four weeks. In contrast, grafts that are heparinized with carbodiimide chemistry exhibit increased intimal hyperplasia (4.2-fold) and increased macrophage infiltration (3.5-fold) compared to ECM-only control grafts. All grafts show similar, partial endothelial cell coverage and little to no ECM remodeling. Overall, a mechanocompatible strategy to improve ECM thromboresistance is described and the importance of ECM mechanical properties for proper in vivo graft performance is highlighted. PMID:27109033

  3. The Biology of the Escherichia coli Extracellular Matrix

    PubMed Central

    Hufnagel, David A.; DePas, William H.; Chapman, Matthew R.

    2015-01-01

    Chapter Summary Escherichia coli (E. coli) is one of the world’s best-characterized organisms, as it has been extensively studied for over a century. However, most of this work has focused on E. coli grown under laboratory conditions that do not faithfully simulate its natural environments. Therefore, the historical perspectives on E. coli physiology and life cycle are somewhat skewed toward experimental systems that feature E. coli growing logarithmically in a test tube. Typically a commensal bacterium, E. coli resides in the lower intestines of a slew of animals. Outside of the lower intestine, E. coli can adapt and survive in a very different set of environmental conditions. Biofilm formation allows E. coli to survive, and even thrive, in environments that do not support the growth of planktonic populations. E. coli can form biofilms virtually everywhere; in the bladder during a urinary tract infection, on in-dwelling medical devices, and outside of the host on plants and in the soil. The E. coli extracellular matrix, primarily composed of the protein polymer named curli and the polysaccharide cellulose, promotes adherence to organic and inorganic surfaces, and resistance to desiccation, the host immune system and other antimicrobials. The pathways that govern E. coli biofilm formation, cellulose production, and curli biogenesis will be discussed in this book chapter, which concludes with insights into the future of E. coli biofilm research and potential therapies. PMID:26185090

  4. The Extracellular Matrix in Bronchopulmonary Dysplasia: Target and Source

    PubMed Central

    Mižíková, Ivana; Morty, Rory E.

    2015-01-01

    Bronchopulmonary dysplasia (BPD) is a common complication of preterm birth that contributes significantly to morbidity and mortality in neonatal intensive care units. BPD results from life-saving interventions, such as mechanical ventilation and oxygen supplementation used to manage preterm infants with acute respiratory failure, which may be complicated by pulmonary infection. The pathogenic pathways driving BPD are not well-delineated but include disturbances to the coordinated action of gene expression, cell–cell communication, physical forces, and cell interactions with the extracellular matrix (ECM), which together guide normal lung development. Efforts to further delineate these pathways have been assisted by the use of animal models of BPD, which rely on infection, injurious mechanical ventilation, or oxygen supplementation, where histopathological features of BPD can be mimicked. Notable among these are perturbations to ECM structures, namely, the organization of the elastin and collagen networks in the developing lung. Dysregulated collagen deposition and disturbed elastin fiber organization are pathological hallmarks of clinical and experimental BPD. Strides have been made in understanding the disturbances to ECM production in the developing lung, but much still remains to be discovered about how ECM maturation and turnover are dysregulated in aberrantly developing lungs. This review aims to inform the reader about the state-of-the-art concerning the ECM in BPD, to highlight the gaps in our knowledge and current controversies, and to suggest directions for future work in this exciting and complex area of lung development (patho)biology. PMID:26779482

  5. Expression of extracellular matrix proteins in adenomatoid odontogenic tumor.

    PubMed

    Modolo, Filipe; Biz, Michelle Tillmann; Martins, Marília Trierveiller; Machado de Sousa, Suzana Orsini; de Araújo, Ney Soares

    2010-03-01

    Altered expression of extracellular matrix (ECM) components has been reported in several pathologies; however, few ECM proteins have been evaluated in adenomatoid odontogenic tumor (AOT). The aim of this study was to analyze the expression and distribution of the ECM proteoglycans: biglycan and decorin; and glycoproteins: osteonectin, osteopontin, bone sialoprotein and osteocalcin in the AOT. Three-micrometer sections from paraffin-embedded specimens were evaluated employing a streptavidin-biotin immunohistochemical method with the antibodies against the proteins previously cited. Only the osteonectin was expressed in the epithelial cells. The eosinophilic amorphous material and the connective tissue showed expression of all components studied. The calcification foci expressed only osteopontin. In conclusion, the low expression of the components studied in neoplastic epithelial cells suggests that the epithelial cells act probably as stimulators of the expression by the stroma, which in turn can act as agonist or antagonist of the tumor growth. These results suggest that the components studied probably have a key role in the biological behavior of the AOT. PMID:20070486

  6. Engineering extracellular matrix structure in 3D multiphase tissues

    PubMed Central

    Gillette, Brian M.; Rossen, Ninna S.; Das, Nikkan; Leong, Debra; Wang, Meixin; Dugar, Arushi; Sia, Samuel K.

    2011-01-01

    In native tissues, microscale variations in the extracellular matrix (ECM) structure can drive different cellular behaviors. Although control over ECM structure could prove useful in tissue engineering and in studies of cellular behavior, isotropic 3D matrices poorly replicate variations in local microenvironments. In this paper, we demonstrate a method to engineer local variations in the density and size of collagen fibers throughout 3D tissues. The results showed that, in engineered multiphase tissues, the structures of collagen fibers in both the bulk ECM phases (as measured by mesh size and width of fibers) as well as at tissue interfaces (as measured by density of fibers and thickness of tissue interfaces) could be modulated by varying the collagen concentrations and gelling temperatures. As the method makes use of a previously published technique for tissue bonding, we also confirmed that significant adhesion strength at tissue interfaces was achieved under all conditions tested. Hence, this study demonstrates how collagen fiber structures can be engineered within all regions of a tightly integrated multiphase tissue scaffold by exploiting knowledge of collagen assembly. PMID:21840047

  7. Modulation of Leukocyte Behavior by an Inflamed Extracellular Matrix

    PubMed Central

    Schor, Hagai; Vaday, Gayle G.

    2000-01-01

    Inflammation is a response of the immune system to foreign insult or physical damage. Various cellular and humoral components of the immune system are recruited from the vascular system and are translocated through endothelium, and into extracellular matrix (ECM) compartments of inflamed tissues. This translocation is orchestrated by various types of accessory signals, in the form of soluble or complexed molecules, which evoke remarkable transitions in leukocyte activities. Recruited inflammatory cells give rise to mechanisms of migration, including the secretion of enzymes and other pro-inflammatory mediators and the alteration of their adhesive contacts with the ECM. Hence, migrating cells secrete enzymes, chemokines, and cytokines which interact with the ECM, and thereby, provide the cells with intrinsic signals for coordinating their responses. Resultant products of enzymatic modifications to the ECM microenvironment, such as cytokine- and ECM-derived molecules, may be also part of a cell-signaling mechanism that provides leukocytes with information about the nature of their inflammatory activity; such a mechanism may give the immune system data that can be cognitively interpreted for consequential activities. This article reviews the findings that support this notion and describe the dynamic interactions between participants of the inflammatory processes. PMID:11097214

  8. Extracellular matrix as an inductive scaffold for functional tissue reconstruction

    PubMed Central

    BROWN, BRYAN N.; BADYLAK, STEPHEN F.

    2014-01-01

    The extracellular matrix (ECM) is a meshwork of both structural and functional proteins assembled in unique tissue-specific architectures. The ECM both provides the mechanical framework for each tissue and organ and is a substrate for cell signaling. The ECM is highly dynamic, and cells both receive signals from the ECM and contribute to its content and organization. This process of “dynamic reciprocity” is key to tissue development and for homeostasis. Based upon these important functions, ECM-based materials have been used in a wide variety of tissue engineering and regenerative medicine approaches to tissue reconstruction. It has been demonstrated that ECM-based materials, when appropriately prepared, can act as inductive templates for constructive remodeling. Specifically, such materials act as templates for the induction of de novo functional, site-appropriate, tissue formation. Herein, the diverse structural and functional roles of the ECM are reviewed to provide a rationale for the use of ECM scaffolds in regenerative medicine. Translational examples of ECM scaffolds in regenerative are provided, and the potential mechanisms by which ECM scaffolds elicit constructive remodeling are discussed. A better understanding of the ability of ECM scaffold materials to define the microenvironment of the injury site will lead to improved clinical outcomes associated with their use. PMID:24291155

  9. Spherical indentation of free-standing acellular extracellular matrix membranes.

    PubMed

    Cloonan, Aidan J; O'Donnell, Michael R; Lee, William T; Walsh, Michael T; De Barra, Eamonn; McGloughlin, Tim M

    2012-01-01

    Numerous scaffold materials have been developed for tissue engineering and regenerative medicine applications to replace or repair damaged tissues and organs. Naturally occurring scaffold materials derived from acellular xenogeneic and autologous extracellular matrix (ECM) are currently in clinical use. These biological scaffold materials possess inherent variations in mechanical properties. Spherical indentation or ball burst testing has commonly been used to evaluate ECM and harvested tissue due to its ease of use and simulation of physiological biaxial loading, but has been limited by complex material deformation profiles. An analytical methodology has been developed and applied to experimental load-deflection data of a model hyperelastic material and lyophilized ECM scaffolds. An optimum rehydration protocol was developed based on water absorption, hydration relaxation and dynamic mechanical analysis. The analytical methodology was compared with finite element simulations of the tests and excellent correlation was seen between the computed biaxial stress resultants and geometry deformations. A minimum rehydration period of 5 min at 37°C was sufficient for the evaluated multilaminated ECM materials. The proposed approach may be implemented for convenient comparative analysis of ECM materials and source tissues, process optimization or during lot release testing. PMID:21864728

  10. The extracellular matrix: Tools and insights for the "omics" era.

    PubMed

    Naba, Alexandra; Clauser, Karl R; Ding, Huiming; Whittaker, Charles A; Carr, Steven A; Hynes, Richard O

    2016-01-01

    The extracellular matrix (ECM) is a fundamental component of multicellular organisms that provides mechanical and chemical cues that orchestrate cellular and tissue organization and functions. Degradation, hyperproduction or alteration of the composition of the ECM cause or accompany numerous pathologies. Thus, a better characterization of ECM composition, metabolism, and biology can lead to the identification of novel prognostic and diagnostic markers and therapeutic opportunities. The development over the last few years of high-throughput ("omics") approaches has considerably accelerated the pace of discovery in life sciences. In this review, we describe new bioinformatic tools and experimental strategies for ECM research, and illustrate how these tools and approaches can be exploited to provide novel insights in our understanding of ECM biology. We also introduce a web platform "the matrisome project" and the database MatrisomeDB that compiles in silico and in vivo data on the matrisome, defined as the ensemble of genes encoding ECM and ECM-associated proteins. Finally, we present a first draft of an ECM atlas built by compiling proteomics data on the ECM composition of 14 different tissues and tumor types. PMID:26163349

  11. Brain extracellular matrix meets COST--matrix for European research networks.

    PubMed

    Gajović, Srećko; Pochet, Roland

    2014-01-01

    Today's researchers are faced with a change from curiosity-driven to mandate-driven research. These two approaches are well combined within scientific networks (Actions) supported by the European Cooperation in Science and Technology (COST) program. The functioning of COST Actions, although directed only to networking, has a substantial impact on European science and can be compared to the functioning of the extracellular matrix in the brain, which although scarce plays a key role in initiation, maintenance, and plasticity of intercellular interactions in the nervous system. COST networks enable interdisciplinary approach and support early-stage researchers, which is a vital asset for the advancement of science. PMID:25410370

  12. The role of cell-extracellular matrix interactions in glomerular injury.

    PubMed

    Borza, Corina M; Pozzi, Ambra

    2012-05-15

    Glomerulosclerosis is characterized by excessive deposition of extracellular matrix within the glomeruli of the kidney, glomerular cell death, and subsequent loss of functional glomeruli. While in physiological situations the levels of extracellular matrix components are kept constant by a tight balance between formation and degradation, in the case of injury that results in fibrosis there is increased matrix deposition relative to its breakdown. Multiple factors control matrix synthesis and degradation, thus contributing to the development of glomerulosclerosis. This review focuses primarily on the role of cell-matrix interactions, which play a critical role in governing glomerular cell cues in both healthy and diseased kidneys. Cell-extracellular matrix interactions are made possible by various cellular receptors including integrins, discoidin domain receptors, and dystroglycan. Upon binding to a selective extracellular matrix protein, these receptors activate intracellular signaling pathways that can either downregulate or upregulate matrix synthesis and deposition. This, together with the observation that changes in the expression levels of matrix receptors have been documented in glomerular disease, clearly emphasizes the contribution of cell-matrix interactions in glomerular injury. Understanding the molecular mechanisms whereby extracellular matrix receptors regulate matrix homeostasis in the course of glomerular injury is therefore critical for devising more effective therapies to treat and ideally prevent glomerulosclerosis. PMID:22417893

  13. THE ROLE OF CELL-EXTRACELLULAR MATRIX INTERACTIONS IN GLOMERULAR INJURY

    PubMed Central

    Borza, Corina M.; Pozzi, Ambra

    2013-01-01

    Glomerulosclerosis is characterized by excessive deposition of extracellular matrix within the glomeruli of the kidney, glomerular cell death, and subsequent loss of functional glomeruli. While in physiological situations the levels of extracellular matrix components are kept constant by a tight balance between formation and degradation, in the case of injury that results in fibrosis there is increased matrix deposition relative to its breakdown. Multiple factors control matrix synthesis and degradation, thus contributing to the development of glomerulosclerosis. This review focuses primarily on the role of cell-matrix interactions, which play a critical role in governing glomerular cell cues in both healthy and diseased kidneys. Cell-extracellular matrix interactions are made possible by various cellular receptors including integrins, discoidin domain receptors, and dystroglycan. Upon binding to a selective extracellular matrix protein, these receptors activate intracellular signaling pathways that can either downregulate or upregulate matrix synthesis and deposition. This, together with the observation that changes in the expression levels of matrix receptors have been documented in glomerular disease, clearly emphasizes the contribution of cell-matrix interactions in glomerular injury. Understanding the molecular mechanisms whereby extracellular matrix receptors regulate matrix homeostasis in the course of glomerular injury is therefore critical for devising more effective therapies to treat and ideally prevent glomerulosclerosis. PMID:22417893

  14. Extracellular Matrix and Dermal Fibroblast Function in the Healing Wound

    PubMed Central

    Tracy, Lauren E.; Minasian, Raquel A.; Caterson, E.J.

    2016-01-01

    Significance: Fibroblasts play a critical role in normal wound healing. Various extracellular matrix (ECM) components, including collagens, fibrin, fibronectin, proteoglycans, glycosaminoglycans, and matricellular proteins, can be considered potent protagonists of fibroblast survival, migration, and metabolism. Recent Advances: Advances in tissue culture, tissue engineering, and ex vivo models have made the examination and precise measurements of ECM components in wound healing possible. Likewise, the development of specific transgenic animal models has created the opportunity to characterize the role of various ECM molecules in healing wounds. In addition, the recent characterization of new ECM molecules, including matricellular proteins, dermatopontin, and FACIT collagens (Fibril-Associated Collagens with Interrupted Triple helices), further demonstrates our cursory knowledge of the ECM in coordinated wound healing. Critical Issues: The manipulation and augmentation of ECM components in the healing wound is emerging in patient care, as demonstrated by the use of acellular dermal matrices, tissue scaffolds, and wound dressings or topical products bearing ECM proteins such as collagen, hyaluronan (HA), or elastin. Once thought of as neutral structural proteins, these molecules are now known to directly influence many aspects of cellular wound healing. Future Directions: The role that ECM molecules, such as CCN2, osteopontin, and secreted protein, acidic and rich in cysteine, play in signaling homing of fibroblast progenitor cells to sites of injury invites future research as we continue investigating the heterotopic origin of certain populations of fibroblasts in a healing wound. Likewise, research into differently sized fragments of the same polymeric ECM molecule is warranted as we learn that fragments of molecules such as HA and tenascin-C can have opposing effects on dermal fibroblasts. PMID:26989578

  15. Customized Interface Biofunctionalization of Decellularized Extracellular Matrix: Toward Enhanced Endothelialization.

    PubMed

    Aubin, Hug; Mas-Moruno, Carlos; Iijima, Makoto; Schütterle, Nicolas; Steinbrink, Meike; Assmann, Alexander; Gil, Francesc Javier; Lichtenberg, Artur; Pegueroles, Marta; Akhyari, Payam

    2016-05-01

    Interface biofunctionalization strategies try to enhance and control the interaction between implants and host organism. Decellularized extracellular matrix (dECM) is widely used as a platform for bioengineering of medical implants, having shown its suitability in a variety of preclinical as well as clinical models. In this study, specifically designed, custom-made synthetic peptides were used to functionalize dECM with different cell adhesive sequences (RGD, REDV, and YIGSR). Effects on in vitro endothelial cell adhesion and in vivo endothelialization were evaluated in standardized models using decellularized ovine pulmonary heart valve cusps (dPVCs) and decellularized aortic grafts (dAoGs), respectively. Contact angle measurements and fluorescent labeling of custom-made peptides showed successful functionalization of dPVCs and dAoGs. The functionalization of dPVCs with a combination of bioactive sequences significantly increased in vitro human umbilical vein endothelial cell adhesion compared to nonfunctionalized controls. In a functional rodent aortic transplantation model, fluorescent-labeled peptides on dAoGs were persistent up to 10 days in vivo under exposure to systemic circulation. Although there was a trend toward enhanced in vivo endothelialization of functionalized grafts compared to nonfunctionalized controls, there was no statistical significance and a large biological variability in both groups. Despite failing to show a clear biological effect in the used in vivo model system, our initial findings do suggest that endothelialization onto dECM may be modulated by customized interface biofunctionalization using the presented method. Since bioactive sequences within the dECM-synthetic peptide platform are easily interchangeable and combinable, further control of host cell proliferation, function, and differentiation seems to be feasible, possibly paving the way to a new generation of multifunctional dECM scaffolds for regenerative medicine. PMID

  16. 3D Extracellular Matrix from Sectioned Human Tissues

    PubMed Central

    Campbell, Catherine B; Cukierman, Edna; Artym, Vira V

    2014-01-01

    Three-dimensional (3D) matrices have significant advantages compared to conventional two-dimensional (2D) matrices for studying cell adhesion, migration, and tissue organization. Cellular behavior is dependent on the surrounding matrix environment for signaling and induction of biological responses (Carletti, et al., 2011; Pampaloni, et al., 2007; Vlodavsky, 1999). 2D cultures induce an artificial polarity in cultured cells between upper and lower surfaces not present normally in the in vivo environment. No longer nonpolar, many aspects of cellular behavior are altered (Beacham, et al., 2007; Grinnell and Petroll, 2010; Yamada and Cukierman, 2007). In addition, 2D models lack the physical properties of 3D matrix, such as topography, stiffness, and dimensionality. To begin to mimic the 3D environment of in vivo connective tissue extracellular matrix (ECM), collagen gels have been used widely (see Unit 10.3). Culture of cells in collagen gels results in a bipolar fibroblast morphology that resembles the in vivo phenotype (Friedl and Brocker, 2000; Even-Ram and Yamada, 2005; Grinnell and Petroll, 2010). Although more physiological, 3D collagen gels lack the complex biochemical and physical microenvironment present in an in vivo ECM that regulates cellular physiological properties (Beacham, et al., 2007). A variety of methods to create a more in vivo-like ECM have been published (Yamada and Cukierman, 2007). Adding critical ECM components to 3D collagen matrices, including fibronectin, hyaluronan, link protein and glycosaminoglycans, can more accurately mimic the structural microenvironment of the native ECM (Friedl and Brocker, 2000). Other ECM models use cultured cell lines, such as fibroblasts, to derive an ECM lattice through secretion of an organized ECM (Beacham, et al., 2007). Different cell lines have been chosen to generate a specific microenvironment for study of particularly types of cellular behavior (Kutys and Yamada, 2013). For example, cultured bovine

  17. Relationships between mechanical properties and extracellular matrix constituents of the cervical stroma during pregnancy.

    PubMed

    House, Michael; Kaplan, David L; Socrate, Simona

    2009-10-01

    In normal pregnancy, the cervix maintains its shape during a period of substantial fetal and uterine growth. Hence, maintenance of biomechanical integrity is an important aspect of cervical function. It is known that cervical mechanical properties arise from extracellular matrix (ECM). The most important constituent of the cervical ECM is fibrillar collagen-it is collagen protein that the cervix derives its "strength" from. Other matrix molecules known to affect the collagen network include water, proteoglycans, hyaluronan, and elastin. The objective of this review is to discuss relationships between biochemical constituents and macroscopic mechanical properties. The individual constituents of the ECM will be discussed, especially in regard to collagen remodeling during pregnancy. In addition, the macroscopic mechanical properties of cervical tissue will be reviewed. An improved understanding of the biochemistry of cervical "strength" will shed light on how the cervix maintains its shape in normal pregnancy and shortens in preterm birth. PMID:19796726

  18. Response of endothelial cells to decellularized extracellular matrix deposited by bone marrow mesenchymal stem cells

    PubMed Central

    Xu, Yue; Yan, Mengdie; Gong, Yihong; Chen, Lei; Zhao, Feng; Zhang, Zhaoqiang

    2014-01-01

    Objective: Evaluate the behavior and function of human umbilical vein endothelial cells (HUVECs) on decellularized extracellular matrix (ECM) deposited by bone marrow mesenchymal stem cells (BMSCs). Methods: Prepared through chemical approach, decellularized ECM was characterized by use of immunofluorescence staining. The morphology, attachment, proliferation and migration of HUVECs cultured on six-well tissue culture plastic (TCP) and decellularized ECM were investigated. Results: Decellularized ECM was successfully prepared without three-dimensional architecture disruption. This biological scaffold is similar to nature vascular ECM, preserved various matrix proteins such as type I collagen, type III collagen and fibronection. HUVECs on decellularized ECM showed well attachment and regular arrangement. Decellularized ECM could also significantly enhance the migration and proliferation potential of HUVECs in contrast to TCP. Conclusion: Deposited by BMSCs, ECM can affect the behavior of endothelial cell and could be used as a promising material in tissue engineering. PMID:25663998

  19. Putative functions of extracellular matrix glycoproteins in secondary palate morphogenesis

    PubMed Central

    d'Amaro, Rocca; Scheidegger, Rolf; Blumer, Susan; Pazera, Pawel; Katsaros, Christos; Graf, Daniel; Chiquet, Matthias

    2012-01-01

    Cleft palate is a common birth defect in humans. Elevation and fusion of paired palatal shelves are coordinated by growth and transcription factors, and mutations in these can cause malformations. Among the effector genes for growth factor signaling are extracellular matrix (ECM) glycoproteins. These provide substrates for cell adhesion (e.g., fibronectin, tenascins), but also regulate growth factor availability (e.g., fibrillins). Cleft palate in Bmp7 null mouse embryos is caused by a delay in palatal shelf elevation. In contrast, palatal shelves of Tgf-β3 knockout mice elevate normally, but a cleft develops due to their failure to fuse. However, nothing is known about a possible functional interaction between specific ECM proteins and Tgf-β/Bmp family members in palatogenesis. To start addressing this question, we studied the mRNA and protein distribution of relevant ECM components during secondary palate development, and compared it to growth factor expression in wildtypewild type and mutant mice. We found that fibrillin-2 (but not fibrillin-1) mRNA appeared in the mesenchyme of elevated palatal shelves adjacent to the midline epithelial cells, which were positive for Tgf-β3 mRNA. Moreover, midline epithelial cells started expressing fibronectin upon contact of the two palatal shelves. These findings support the hypothesis that fibrillin-2 and fibronectin are involved in regulating the activity of Tgf-β3 at the fusing midline. In addition, we observed that tenascin-W (but not tenascin-C) was misexpressed in palatal shelves of Bmp7-deficient mouse embryos. In contrast to tenascin-C, tenascin-W secretion was strongly induced by Bmp7 in embryonic cranial fibroblasts in vitro. These results are consistent with a putative function for tenascin-W as a target of Bmp7 signaling during palate elevation. Our results indicate that distinct ECM proteins are important for morphogenesis of the secondary palate, both as downstream effectors and as regulators of Tgf

  20. Putative functions of extracellular matrix glycoproteins in secondary palate morphogenesis.

    PubMed

    d'Amaro, Rocca; Scheidegger, Rolf; Blumer, Susan; Pazera, Pawel; Katsaros, Christos; Graf, Daniel; Chiquet, Matthias

    2012-01-01

    Cleft palate is a common birth defect in humans. Elevation and fusion of paired palatal shelves are coordinated by growth and transcription factors, and mutations in these can cause malformations. Among the effector genes for growth factor signaling are extracellular matrix (ECM) glycoproteins. These provide substrates for cell adhesion (e.g., fibronectin, tenascins), but also regulate growth factor availability (e.g., fibrillins). Cleft palate in Bmp7 null mouse embryos is caused by a delay in palatal shelf elevation. In contrast, palatal shelves of Tgf-β3 knockout mice elevate normally, but a cleft develops due to their failure to fuse. However, nothing is known about a possible functional interaction between specific ECM proteins and Tgf-β/Bmp family members in palatogenesis. To start addressing this question, we studied the mRNA and protein distribution of relevant ECM components during secondary palate development, and compared it to growth factor expression in wildtypewild type and mutant mice. We found that fibrillin-2 (but not fibrillin-1) mRNA appeared in the mesenchyme of elevated palatal shelves adjacent to the midline epithelial cells, which were positive for Tgf-β3 mRNA. Moreover, midline epithelial cells started expressing fibronectin upon contact of the two palatal shelves. These findings support the hypothesis that fibrillin-2 and fibronectin are involved in regulating the activity of Tgf-β3 at the fusing midline. In addition, we observed that tenascin-W (but not tenascin-C) was misexpressed in palatal shelves of Bmp7-deficient mouse embryos. In contrast to tenascin-C, tenascin-W secretion was strongly induced by Bmp7 in embryonic cranial fibroblasts in vitro. These results are consistent with a putative function for tenascin-W as a target of Bmp7 signaling during palate elevation. Our results indicate that distinct ECM proteins are important for morphogenesis of the secondary palate, both as downstream effectors and as regulators of Tgf

  1. Extracellular matrix remodelling in response to venous hypertension: proteomics of human varicose veins

    PubMed Central

    Barallobre-Barreiro, Javier; Oklu, Rahmi; Lynch, Marc; Fava, Marika; Baig, Ferheen; Yin, Xiaoke; Barwari, Temo; Potier, David N.; Albadawi, Hassan; Jahangiri, Marjan; Porter, Karen E.; Watkins, Michael T.; Misra, Sanjay; Stoughton, Julianne; Mayr, Manuel

    2016-01-01

    Aims Extracellular matrix remodelling has been implicated in a number of vascular conditions, including venous hypertension and varicose veins. However, to date, no systematic analysis of matrix remodelling in human veins has been performed. Methods and results To understand the consequences of venous hypertension, normal and varicose veins were evaluated using proteomics approaches targeting the extracellular matrix. Varicose saphenous veins removed during phlebectomy and normal saphenous veins obtained during coronary artery bypass surgery were collected for proteomics analysis. Extracellular matrix proteins were enriched from venous tissues. The proteomics analysis revealed the presence of >150 extracellular matrix proteins, of which 48 had not been previously detected in venous tissue. Extracellular matrix remodelling in varicose veins was characterized by a loss of aggrecan and several small leucine-rich proteoglycans and a compensatory increase in collagen I and laminins. Gene expression analysis of the same tissues suggested that the remodelling process associated with venous hypertension predominantly occurs at the protein rather than the transcript level. The loss of aggrecan in varicose veins was paralleled by a reduced expression of aggrecanases. Chymase and tryptase β1 were among the up-regulated proteases. The effect of these serine proteases on the venous extracellular matrix was further explored by incubating normal saphenous veins with recombinant enzymes. Proteomics analysis revealed extensive extracellular matrix degradation after digestion with tryptase β1. In comparison, chymase was less potent and degraded predominantly basement membrane-associated proteins. Conclusion The present proteomics study provides unprecedented insights into the expression and degradation of structural and regulatory components of the vascular extracellular matrix in varicosis. PMID:27068509

  2. Targeted rehabilitation after extracellular matrix scaffold transplantation for the treatment of volumetric muscle loss.

    PubMed

    Gentile, Natalie E; Stearns, Kristen M; Brown, Elke H P; Rubin, J Peter; Boninger, Michael L; Dearth, Christopher L; Ambrosio, Fabrisia; Badylak, Stephen F

    2014-11-01

    Rehabilitation therapy is an important aspect of recovery after volumetric muscle loss. However, the traditional rehabilitation approach involves a period of rest and passive loading followed by gradual active loading. Extracellular matrix is a naturally occurring material consisting of structural proteins that provide mechanical strength, structural support, and functional molecules with diverse bioactive properties. There is evidence to suggest that the addition of aggressive regenerative rehabilitation protocols immediately after surgical implantation of an extracellular matrix scaffold to an area of volumetric muscle loss has significant benefits for extracellular matrix remodeling. Rehabilitation exercises likely provide the needed mechanical signals to encourage cell migration and site-specific differentiation in the temporal framework required for constructive remodeling. Herein, the authors review the literature and present an example of an aggressive rehabilitation program implemented immediately after extracellular matrix transplantation into a severely injured quadriceps muscle. PMID:25133624

  3. Glycation of extracellular matrix proteins impairs migration of immune cells.

    PubMed

    Haucke, Elisa; Navarrete-Santos, Alexander; Simm, Andreas; Silber, Rolf-Edgar; Hofmann, Britt

    2014-01-01

    The immune response during aging and diabetes is disturbed and may be due to the altered migration of immune cells in an aged tissue. Our study should prove the hypothesis that age and diabetes-related advanced glycation end products (AGEs) have an impact on the migration and adhesion of human T-cells. To achieve our purpose, we used in vitro AGE-modified proteins (soluble albumin and fibronectin [FN]), as well as human collagen obtained from bypass graft. A Boyden chamber was used to study cell migration. Migrated Jurkat T-cells were analyzed by flow cytometry and cell adhesion by crystal violet staining. Actin polymerization was determined by phalloidin-Alexa-fluor 488-labeled antibody and fluorescence microscopy. We found that significantly fewer cells (50%, p = 0.003) migrated through methylglyoxal modified FN. The attachment to FN in the presence of AGE-bovine serum albumin (BSA) was also reduced (p < 0.05). In ex vivo experiments, isolated collagen from human vein graft material negatively affected the migration of the cells depending on the grade of AGE modification of the collagen. Collagen with a low AGE level reduced the cell migration by 30%, and collagen with a high AGE level by 60%. Interaction of the cells with an AGE-modified matrix, but not with soluble AGEs like BSA-AGE per se, was responsible for a disturbed migration. The reduced migration was accompanied by an impaired actin polymerization. We conclude that AGEs-modified matrix protein inhibits cell migration and adhesion of Jurkat T-cells. PMID:24635174

  4. [Disorders of the extracellular matrix in epithelial-stromal and stromal corneal dystrophies].

    PubMed

    Varkoly, Gréta; Bencze, János; Módis, László; Hortobágyi, Tibor

    2016-08-01

    The human cornea is rich in extracellular matrix. The stroma constitutes the main thickness of the cornea, which consists of collagens and proteoglycans mainly. The epithelial-stromal and stromal dystrophies of the cornea are either autosomal dominant or recessive inherited disorders, which are unrelated to inflammation or trauma. The diseases can manifest in each layer of the cornea, but in most cases the corneal stroma is affected. Generally, they develop in childhood or young adulthood but the diagnosis is only possible when clinical signs (epithelial erosions, decreased visual acuity, photophobia) develop. The different protein aggregates (hyaline, amyloid, crystalline) deposited in the corneal layers result in mild or advanced corneal opacity and loss of the corneal transparency due to disorganisation of the extracellular matrix. In some of the corneal dystrophies the keratane sulphate proteoglycan looses its function which results in a loss of the regular interfibrillar spacing. Due to the severe corneal opacity patients may need corneal transplantation. Orv. Hetil., 2016, 157(33), 1299-1303. PMID:27523312

  5. Extracellular Matrix Components Regulate Cellular Polarity and Tissue Structure in the Developing and Mature Retina

    PubMed Central

    Varshney, Shweta; Hunter, Dale D.; Brunken, William J.

    2015-01-01

    While genetic networks and other intrinsic mechanisms regulate much of retinal development, interactions with the extracellular environment shape these networks and modify their output. The present review has focused on the role of one family of extracellular matrix molecules and their signaling pathways in retinal development. In addition to their effects on the developing retina, laminins play a role in maintaining Müller cell polarity and compartmentalization, thereby contributing to retinal homeostasis. This article which is intended for the clinical audience, reviews the fundamentals of retinal development, extracellular matrix organization and the role of laminins in retinal development. The role of laminin in cortical development is also briefly discussed. PMID:26730321

  6. Illustrating the interplay between the extracellular matrix and microRNAs

    PubMed Central

    Piccinini, Anna M; Midwood, Kim S

    2014-01-01

    The discovery of cell surface receptors that bind to extracellular matrix (ECM) components marked a new era in biological research. Since then there has been an increasing appreciation of the importance of studying cells in the context of their extracellular environment. Cell behaviour is profoundly affected by the ECM, whose synthesis and turnover must be finely balanced in order to maintain normal function and prevent disease. In the last decade, microRNAs (miRNAs) have emerged as key regulators of ECM gene expression. As new technologies for the identification and validation of miRNA targets continue to be developed, a growing body of data supporting the role of miRNAs in regulating the ECM biology has arisen from a variety of cell and animal models along with clinical studies. However, more recent findings suggest an intriguing interplay between the ECM and miRNAs: not only can miRNAs control the composition of the ECM, but also the ECM can affect the expression of specific miRNAs. Here we discuss how miRNAs contribute to the synthesis, maintenance and remodelling of the ECM during development and disease. Furthermore, we bring to light evidence that points to a role for the ECM in regulating miRNA expression and function. PMID:24761792

  7. Cartilage extracellular matrix metabolism differs in serum and synovial fluid.

    PubMed

    Martin, James A; Wilkey, Andrew L; Brand, Richard A

    2002-01-01

    Most cartilage explant culture studies assume conventional serum-supplemented growth media are biologically equivalent to the natural synovial fluid which baths cartilage in vivo. Few studies have systematically compared the effects of serum versus synovial fluid in culture. To address this assumption we conducted a series of studies to determine if cartilage matrix synthesis is significantly different in serum-based versus synovial fluid-based media. Normal bovine cartilage explants were cultured in DMEM either alone or supplemented with bovine serum or bovine synovial fluid. Matrix synthesis was measured with radiolabeling techniques. We then compared responses to insulin-like growth factor I (IGF-I, a stimulator of matrix synthesis), and interleukin-1beta (IL-1beta, an inhibitor of matrix synthesis). We observed significantly lower matrix synthesis activity in synovial fluid versus serum. Caution shoud be used in extrapolating studies of cartilage grown in media supplemented with serum rather than synovial fluid. PMID:12843702

  8. Elasticity-mediated nematiclike bacterial organization in model extracellular DNA matrix.

    PubMed

    Smalyukh, Ivan I; Butler, John; Shrout, Joshua D; Parsek, Matthew R; Wong, Gerard C L

    2008-09-01

    DNA is a common extracellular matrix component of bacterial biofilms. We find that bacteria can spontaneously order in a matrix of aligned concentrated DNA, in which rod-shaped cells of Pseudomonas aeruginosa follow the orientation of extended DNA chains. The alignment of bacteria is ensured by elasticity and liquid crystalline properties of the DNA matrix. These findings show how behavior of planktonic bacteria may be modified in extracellular polymeric substances of biofilms and illustrate the potential of using complex fluids to manipulate embedded nanosized and microsized active particles. PMID:18850984

  9. The extracellular matrix and blood vessel formation: not just a scaffold

    PubMed Central

    Rhodes, John M; Simons, Michael

    2007-01-01

    Abstract The extracellular matrix plays a number of important roles, among them providing structural support and information to cellular structures such as blood vessels imbedded within it. As more complex organisms have evolved, the matrix ability to direct signalling towards the vasculature and remodel in response to signalling from the vasculature has assumed progressively greater importance. This review will focus on the molecules of the extracellular matrix, specifically relating to vessel formation and their ability to signal to the surrounding cells to initiate or terminate processes involved in blood vessel formation. PMID:17488472

  10. An investigation of the influence of extracellular matrix anisotropy and cell-matrix interactions on tissue architecture.

    PubMed

    Dyson, R J; Green, J E F; Whiteley, J P; Byrne, H M

    2016-06-01

    Mechanical interactions between cells and the fibrous extracellular matrix (ECM) in which they reside play a key role in tissue development. Mechanical cues from the environment (such as stress, strain and fibre orientation) regulate a range of cell behaviours, including proliferation, differentiation and motility. In turn, the ECM structure is affected by cells exerting forces on the matrix which result in deformation and fibre realignment. In this paper we develop a mathematical model to investigate this mechanical feedback between cells and the ECM. We consider a three-phase mixture of collagen, culture medium and cells, and formulate a system of partial differential equations which represents conservation of mass and momentum for each phase. This modelling framework takes into account the anisotropic mechanical properties of the collagen gel arising from its fibrous microstructure. We also propose a cell-collagen interaction force which depends upon fibre orientation and collagen density. We use a combination of numerical and analytical techniques to study the influence of cell-ECM interactions on pattern formation in tissues. Our results illustrate the wide range of structures which may be formed, and how those that emerge depend upon the importance of cell-ECM interactions. PMID:26328534

  11. Smooth muscle cell expression of extracellular matrix genes after arterial injury.

    PubMed Central

    Nikkari, S. T.; Järveläinen, H. T.; Wight, T. N.; Ferguson, M.; Clowes, A. W.

    1994-01-01

    Accumulation of extracellular matrix (ECM) after arterial injury is an important event in the development of intimal thickening and is modulated by heparin. To investigate the regulation of matrix protein expression, we have analyzed messenger RNA levels by Northern blotting for various ECM proteins in the rat carotid artery balloon injury model. RNA was extracted from normal arteries and from intima-medial preparations at 2 days, 1 week, 2 weeks, and 4 weeks after balloon injury of arteries in animals receiving either saline or heparin infusion. Transcripts for the heparan sulfate proteoglycans perlecan, syndecan, and ryudocan; the chondroitin sulfate proteoglycan versican; the dermatan sulfate proteoglycan biglycan; type I procollagen; and tropoelastin all were increased on Northern blots beginning at 1 week after injury. By in situ hybridization, the transcripts for elastin nd biglycan were primarily localized to smooth muscle cells in the intima and were diminished by heparin in proportion to the decrease in intimal mass. Other matrix genes (perlecan, ryudocan) were expressed in the intima and media and were not affected by heparin. The results support the conclusion that ECM gene expression is a relatively late event in the response of the carotid artery, and that some of the genes are expressed only in the intima whereas others are expressed in both the intima and media. Images Figure 2 Figure 3 Figure 4 PMID:8203472

  12. Distribution of extracellular matrix components in normal and degenerated canine tricuspid valve leaflets.

    PubMed

    Aupperle, H; März, I; Thielebein, J; Kiefer, B; Dinges, G; Schoon, H-A

    2009-07-01

    The aim of the present study was to investigate the composition and distribution of various extracellular matrix (ECM) components in normal canine tricuspid valves (TVs) and in TVs affected by chronic valvular disease (CVD). The parietal (pTV) and septal (sTV) leaflets of the TVs from 27 dogs were investigated immunohistochemically for expression of collagen types I, III, IV and VI, elastin, laminin, fibronectin and heparan sulphate. Normal pTV consisted mainly of elastin and collagen VI in the atrialis, fibronectin in the thin spongiosa and mixed collagens in the fibrosa. The layered structure was less distinct in sTV, with numerous adipocytes and proteoglycans in the spongiosa and collagen III predominating in the fibrosa. The earliest stages of CVD affecting the pTV were recognized in the spongiosa and progression to advanced disease was characterized by nodular accumulation of proteoglycans within the free edge of the leaflet. These nodular lesions of the pTV contained more fibronectin, elastin and collagens I and VI than those affecting the sTV. These findings contrast with those reported in CVD affecting the mitral valve (MV) in which the early lesions affect the atrialis and advanced disease involves the entire leaflet. The pathogenesis of CVD in TV may involve initial alterations of the tricuspid annulus that lead to early lesions within the spongiosa, resulting in further shear stress and proteoglycan accumulation at the free edge of the pTV. PMID:19342061

  13. Extracellular matrix density regulates the rate of neovessel growth and branching in sprouting angiogenesis.

    PubMed

    Edgar, Lowell T; Underwood, Clayton J; Guilkey, James E; Hoying, James B; Weiss, Jeffrey A

    2014-01-01

    Angiogenesis is regulated by the local microenvironment, including the mechanical interactions between neovessel sprouts and the extracellular matrix (ECM). However, the mechanisms controlling the relationship of mechanical and biophysical properties of the ECM to neovessel growth during sprouting angiogenesis are just beginning to be understood. In this research, we characterized the relationship between matrix density and microvascular topology in an in vitro 3D organ culture model of sprouting angiogenesis. We used these results to design and calibrate a computational growth model to demonstrate how changes in individual neovessel behavior produce the changes in vascular topology that were observed experimentally. Vascularized gels with higher collagen densities produced neovasculatures with shorter vessel lengths, less branch points, and reduced network interconnectivity. The computational model was able to predict these experimental results by scaling the rates of neovessel growth and branching according to local matrix density. As a final demonstration of utility of the modeling framework, we used our growth model to predict several scenarios of practical interest that could not be investigated experimentally using the organ culture model. Increasing the density of the ECM significantly reduced angiogenesis and network formation within a 3D organ culture model of angiogenesis. Increasing the density of the matrix increases the stiffness of the ECM, changing how neovessels are able to deform and remodel their surroundings. The computational framework outlined in this study was capable of predicting this observed experimental behavior by adjusting neovessel growth rate and branching probability according to local ECM density, demonstrating that altering the stiffness of the ECM via increasing matrix density affects neovessel behavior, thereby regulated vascular topology during angiogenesis. PMID:24465500

  14. The Matrix Reloaded: How Sensing the Extracellular Matrix Synchronizes Bacterial Communities

    PubMed Central

    Steinberg, Nitai

    2015-01-01

    In response to chemical communication, bacterial cells often organize themselves into complex multicellular communities that carry out specialized tasks. These communities are frequently referred to as biofilms, which involve the collective behavior of different cell types. Like cells of multicellular eukaryotes, the biofilm cells are surrounded by self-produced polymers that constitute the extracellular matrix (ECM), which binds them to each other and to the surface. In multicellular eukaryotes, it has been evident for decades that cell-ECM interactions control multiple cellular processes during development. While cells both in biofilms and in multicellular eukaryotes are surrounded by ECM and activate various genetic programs, until recently it has been unclear whether cell-ECM interactions are recruited in bacterial communicative behaviors. In this review, we describe the examples reported thus far for ECM involvement in control of cell behavior throughout the different stages of biofilm formation. The studies presented in this review have provided a newly emerging perspective of the bacterial ECM as an active player in regulation of biofilm development. PMID:25825428

  15. Extracellular Matrix and Regenerative Therapies from the Cardiac Perspective.

    PubMed

    Dogan, Arin; Parmaksız, Mahmut; Elçin, A Eser; Elçin, Y Murat

    2016-04-01

    Cardiovascular diseases are the leading cause of death and a major cause of financial burden. Regenerative therapies for heart diseases bring the promise of alternative treatment modalities for myocardial infarction, ischemic heart disease, and congestive heart failure. Although, clinical trials attest to the safety of stem cell injection therapies, researchers need to overcome the underlying mechanisms that are limiting the success of future regenerative options. This article aims to review the basic scientific concepts in the field of mechanobiology and the effects of extracellular functions on stem cell fate. PMID:26668014

  16. Global transcriptome analysis of Human Bone Marrow Stromal Cells (BMSCs) reveals proliferative, mobile, and Interactive cells that produce abundant extracellular matrix proteins, some of which may affect BMSC Potency

    PubMed Central

    Ren, Jiaqiang; Jin, Ping; Sabatino, Marianna; Balakumaran, Arun; Feng, Ji; Kuznetsov, Sergei A.; Klein, Harvey G.; Robey, Pamela G.; Stroncek, David F.

    2012-01-01

    Background Bone marrow stromal cells (BMSCs) are being used for immune modulatory, anti-inflammatory and tissue engineering applications, but the properties responsible for these effects are not completely understood. Human BMSCs were characterized to identify factors that might be responsible for their clinical effects and biomarkers for assessing their quality. Methods Early passage BMSCs prepared from marrow aspirates of 4 healthy subjects were compared to 3 human embryonic stem cell (hESC) samples, CD34+ cells from 3 healthy subjects and 3 fibroblast cell lines. The cells were analyzed with oligonucleotide expression microarrays with more than 35,000 probes. Results BMSC gene expression signatures of BMSCs differed from those of hematopoietic stem cells (HSCs), hESCs and fibroblasts. Genes up-regulated in BMSCs were involved with cell movement, cell-to-cell signaling and interaction and proliferation. The up-regulated genes most likely belonged to pathways for integrin signaling, integrin linked kinase (ILK) signaling, NFR2-mediated oxidative stress response, regulation of actin-based motility by Rho, actin cytoskeletal signaling, caveolar-mediated endocytosis, clathrin-mediated endocytosis and Wnt/β catenin signaling. Among the most highly up-regulated genes were structural extracellular (ECM) proteins:α5 and β 5 integrin chains, fibronectin, collagen type IIIα1 and Vα1; and functional EMC proteins: connective tissue growth factor (CTGF), transforming growth factor beta induced protein (TGFBI) and ADAM12. Conclusions Global analysis of human BMSCs suggests that they are mobile, metabolically active, proliferative and interactive cells that make use of integrins and integrin signaling. They produce abundant ECM proteins that may contribute to their clinical immune modulatory and anti-inflammatory effects. PMID:21250865

  17. A potential role for glia-derived extracellular matrix remodeling in postinjury epilepsy.

    PubMed

    Kim, Soo Young; Porter, Brenda E; Friedman, Alon; Kaufer, Daniela

    2016-09-01

    Head trauma and vascular injuries are known risk factors for acquired epilepsy. The sequence of events that lead from the initial injury to the development of epilepsy involves complex plastic changes and circuit rewiring. In-depth, comprehensive understanding of the epileptogenic process is critical for the identification of disease-modifying targets. Here we review the complex interactions of cellular and extracellular components that may promote epileptogenesis, with an emphasis on the role of astrocytes. Emerging evidence demonstrates that astrocytes promptly respond to brain damage and play a critical role in the development of postinjury epilepsy. Astrocytes have been shown to regulate extracellular matrix (ECM) remodeling, which can affect plasticity and stability of synapses and, in turn, contribute to the epileptogenic process. From these separate lines of evidence, we present a hypothesis suggesting a possible role for astrocyte-regulated remodeling of ECM and perineuronal nets, a specialized ECM structure around fast-spiking inhibitory interneurons, in the development and progression of posttraumatic epilepsies. © 2016 Wiley Periodicals, Inc. PMID:27265805

  18. Formation of atypical podosomes in extravillous trophoblasts regulates extracellular matrix degradation

    PubMed Central

    Patel, Anand; Dash, Philip R.

    2012-01-01

    Throughout pregnancy the cytotrophoblast, the stem cell of the placenta, gives rise to the differentiated forms of trophoblasts. The two main cell lineages are the syncytiotrophoblast and the invading extravillous trophoblast. A successful pregnancy requires extravillous trophoblasts to migrate and invade through the decidua and then remodel the maternal spiral arteries. Many invasive cells use specialised cellular structures called invadopodia or podosomes in order to degrade extracellular matrix. Despite being highly invasive cells, the presence of invadapodia or podosomes has not previously been investigated in trophoblasts. In this study these structures have been identified and characterised in extravillous trophoblasts. The role of specialised invasive structures in trophoblasts in the degradation of the extracellular matrix was compared with well characterised podosomes and invadopodia in other invasive cells and the trophoblast specific structures were characterised by using a sensitive matrix degradation assay which enabled visualisation of the structures and their dynamics. We show trophoblasts form actin rich protrusive structures which have the ability to degrade the extracellular matrix during invasion. The degradation ability and dynamics of the structures closely resemble podosomes, but have unique characteristics that have not previously been described in other cell types. The composition of these structures does not conform to the classic podosome structure, with no distinct ring of plaque proteins such as paxillin or vinculin. In addition, trophoblast podosomes protrude more deeply into the extracellular matrix than established podosomes, resembling invadopodia in this regard. We also show several significant pathways such as Src kinase, MAPK kinase and PKC along with MMP-2 and 9 as key regulators of extracellular matrix degradation activity in trophoblasts, while podosome activity was regulated by the rigidity of the extracellular matrix. PMID

  19. Cell density modulates growth, extracellular matrix, and protein synthesis of cultured rat mesangial cells.

    PubMed

    Wolthuis, A; Boes, A; Grond, J

    1993-10-01

    Mesangial cell (MC) hyperplasia and accumulation of extracellular matrix are hallmarks of chronic glomerular disease. The present in vitro study examined the effects of cell density on growth, extracellular matrix formation, and protein synthesis of cultured rat MCs. A negative linear relationship was found between initial plating density and DNA synthesis per cell after 24 hours incubation in medium with 10% fetal calf serum (range: 1 x 10(3) to 7 x 10(5) MCs/2cm2, r = 0.996, P < 0.001). Enzyme-linked immunosorbent assay of the amount of fibronectin in the conditioned medium after 72 hours showed a negative relationship with increasing cell density. In contrast, the amount of cell-associated fibronectin increased to maximal values in confluent cultures, and no further increase was seen at supraconfluency. The relative collagen synthesis in the conditioned medium and cell layer--assessed by collagenase digestion after 5 hours [3H]proline pulse labeling--showed a similar pattern. Secreted collagen decreased with increasing cell density from 3.4% to 0.2% of total protein synthesis. In contrast, cell-associated collagen increased from 1.1% to 11.8% of newly synthesized protein until confluency followed by a decrease to 4.2% at supraconfluency. Specific immunoprecipitation of collagen types I, III, and IV revealed a significant (twofold) increase in collagen I synthesis per cell at confluency. Collagen III and IV synthesis was not affected by cell density. Specific protein expression in both the medium and cell layer were analyzed by two-dimensional polyacrylamide gel electrophoresis (150 to 20 kd, pI 5.0 to 7.0) after 20 hours steady-state metabolic labeling with [35S]methionine. Supraconfluent MCs displayed overexpression of 10, underexpression of four, new expression of five, and changed mobility of three different intracellular proteins. Of interest was the overexpression of two proteins (89 kd, pI 5.31 and 72 kd, pI 5.32) that were identified by immunoblotting as

  20. Tissue architecture and breast cancer: the role of extracellular matrix and steroid hormones

    SciTech Connect

    Hansen, R K; Bissell, M J

    2000-06-01

    The changes in tissue architecture that accompany the development of breast cancer have been the focus of investigations aimed at developing new cancer therapeutics. As we learn more about the normal mammary gland, we have begun to understand the complex signaling pathways underlying the dramatic shifts in the structure and function of breast tissue. Integrin-, growth factor-, and steroid hormone-signaling pathways all play an important part in maintaining tissue architecture; disruption of the delicate balance of signaling results in dramatic changes in the way cells interact with each other and with the extracellular matrix, leading to breast cancer. The extracellular matrix itself plays a central role in coordinating these signaling processes. In this review, we consider the interrelationships between the extracellular matrix, integrins, growth factors, and steroid hormones in mammary gland development and function.

  1. Responses of cultured neural retinal cells to substratum-bound laminin and other extracellular matrix molecules.

    PubMed

    Adler, R; Jerdan, J; Hewitt, A T

    1985-11-01

    The responses of cultured chick embryo retinal neurons to several extracellular matrix molecules are described. Retinal cell suspensions in serum-free medium containing the "N1" supplement (J. E. Bottenstein, S. D. Skaper, S. Varon, and J. Sato, 1980, Exp. Cell Res. 125, 183-190) were seeded on tissue culture plastic surfaces pretreated with polyornithine (PORN) and with one of the factors to be tested. Substantial cell survival could be observed after 72 hr in vitro on PORN pretreated with serum or laminin, whereas most cells appeared to be degenerating on untreated PORN, PORN-fibronectin, and PORN-chondronectin. Cell attachment, although quantitatively similar for all these substrata, was temperature-dependent on serum and laminin but not on fibronectin or untreated PORN. In a short-term bioassay, neurite development was abundant on laminin, scarce on serum and fibronectin, and absent on PORN. No positive correlation between cell spreading and neurite production could be seen: cell spreading was more extensive on PORN and fibronectin than on laminin or serum, while on laminin-treated dishes, spreading was similar for neurite-bearing and non-neurite-bearing cells. Laminin effects on retinal neurons were clearly substratum dependent. When bound to tissue culture plastic, laminin showed a dose-dependent inhibitory effect on cell attachment and did not stimulate neurite development. PORN-bound laminin, on the other hand, did not affect cell attachment but caused marked stimulation of neurite development, suggesting that laminin conformation and/or the spatial distribution of active sites play an important role in the neurite-promoting function of this extracellular matrix molecule. Investigation of the embryonic retina with ELISA and immunocytochemical methods showed that laminin is present in this organ during development. Therefore, in vivo and in vitro observations are consistent with the possibility that laminin might influence neuronal development in the retina

  2. The role of extracellular matrix in lateral transmission of force in skeletal muscle

    NASA Astrophysics Data System (ADS)

    Gao, Yingxin

    This dissertation describes the role of extracellular matrix (ECM) in the lateral transmission of force. It consists of an experimental studies of the ECM and mathematical modeling of lateral transmission of force. The effect of aging on the structural and mechanical properties of the epimysium of muscle of the rats were examined. No statistically significant differences were found in the ultrastructure, or the thickness of the epimysium. However, from the tensile stress-strain tests, it was found that the epimysium of muscles from old rats was much stiffer than that of the young rats. Based on these observations. It was concluded that the differences in the mechanical properties of the epimysium of the muscles from the old compared with young rats were not associated with the arrangement and size of collagen fibers in the epimysium. Consequently, other methods will be required to identify the structural bases of the mechanical differences. The stress-strain relationships for the epimysiums of the skeletal muscles from both the young and old rats were found to be nonlinear. A mathematical model was developed that showed that the nonlinear behavior results from the waviness and the reorientation of the collagen fibers in the epimysium. The ECM plays an important role in lateral transmission of force in skeletal muscle by providing shear stress between the muscle fibers or fascicles. A mathematical model was developed to investigate the mechanisms of lateral transmission. It was a modification of the shear lag theory for chopped fiber composite materials used in engineering applications. The modified shear lag theory includes an activation strain to account for muscle contraction and a myofibrils-endomysium interfaces that accounts for the molecular lateral linkages. The model was used to simulate the classic experiments of Street. It was demonstrated that lateral transmission of force in the skeletal muscle is affected by the mechanical and structural properties of

  3. Regulation of Extracellular Matrix Organization by BMP Signaling in Caenorhabditis elegans

    PubMed Central

    Schultz, Robbie D.; Bennett, Emily E.; Ellis, E. Ann; Gumienny, Tina L.

    2014-01-01

    In mammals, Bone Morphogenetic Protein (BMP) pathway signaling is important for the growth and homeostasis of extracellular matrix, including basement membrane remodeling, scarring, and bone growth. A conserved BMP member in Caenorhabditis elegans, DBL-1, regulates body length in a dose-sensitive manner. Loss of DBL-1 pathway signaling also results in increased anesthetic sensitivity. However, the physiological basis of these pleiotropic phenotypes is largely unknown. We created a DBL-1 over-expressing strain and show that sensitivity to anesthetics is inversely related to the dose of DBL-1. Using pharmacological, genetic analyses, and a novel dye permeability assay for live, microwave-treated animals, we confirm that DBL-1 is required for the barrier function of the cuticle, a specialized extracellular matrix. We show that DBL-1 signaling is required to prevent animals from forming tail-entangled aggregates in liquid. Stripping lipids off the surface of wild-type animals recapitulates this phenotype. Finally, we find that DBL-1 signaling affects ultrastructure of the nematode cuticle in a dose-dependent manner, as surface lipid content and cuticular organization are disrupted in animals with genetically altered DBL-1 levels. We propose that the lipid layer coating the nematode cuticle normally prevents tail entanglement, and that reduction of this layer by loss of DBL-1 signaling promotes aggregation. This work provides a physiological mechanism that unites the DBL-1 signaling pathway roles of not only body size regulation and drug responsiveness, but also the novel Hoechst 33342 staining and aggregation phenotypes, through barrier function, content, and organization of the cuticle. PMID:25013968

  4. Symposium: Role of the extracellular matrix in mammary development. Regulation of milk protein and basement membrane gene expression: The influence of the extracellular matrix

    SciTech Connect

    Aggeler, J.; Park, C.S.; Bissell, M.J.

    1988-10-01

    Synthesis and secretion of milk proteins ({alpha}-casein, {beta}-casein, {gamma}-casein, and transferrin) by cultured primary mouse mammary epithelial cells is modulated by the extracellular matrix. In cells grown on released or floating type I collagen gels, mRNA for {beta}-casein and transferrin is increased as much as 30-fold over cells grown on plastic. Induction of {beta}-casein expression depends strongly on the presence of lactogenic hormones, especially prolactin, in the culture. When cells are plated onto partially purified reconstituted basement membrane, dramatic changes in morphology and milk protein gene expression are observed. Cells cultured on the matrix for 6 to 8 d in the presence of prolactin, insulin, and hydrocortisone form hollow spheres and duct-like structures that are completely surrounded by matrix. The cells lining these spheres appear actively secretory and are oriented with their apices facing the lumen. Hybridization experiments indicate that mRNA for {beta}-casein can be increased as much as 70-fold in these cultures. Because > 90% of the cultured cells synthesize immunoreactive {beta}-casein, as compared with only 40% of cells in the late pregnant gland, the matrix appears to be able to induce protein expression in previously silent cells. Synthesis of laminin and assembly of a mammary-specific basal lamina by cells cultured on different extracellular matrices also appears to depend on the presence of lactogenic hormones. These studies provide support for the concept of dynamic reciprocity in which complex interactions between extracellular matrix and the cellular cytoskeleton contribute to the induction and maintenance of tissue-specific gene expression in the mammary gland.

  5. [Effect of fibronectin on the synthesis of extracellular matrix proteins in periodontal ligament cells].

    PubMed

    Wan, L; Wu, Z; Zhou, Y

    1996-11-01

    Immunofluorescence staining method and fluorescence spectrophotometry were used to study the synthesis of extracellular matrix proteins in periodontal ligament cells (PDL cells) when exogenous fibronectin (FN) existed. The results showed that the right amount of exogenous FN (0.044 mumol/l) could increase the amount of type I collagen and type III collagen in PDL cells (P < 0.01), inhibit the synthesis of FN itself (P < 0.01). It suggested that exogenous FN can effect the synthesis of extracellular matrix proteins so as to promote a new connective tissue attachment formation. PMID:9592289

  6. Depressed immune surveillance against cancer: role of deficient T cell: extracellular matrix interactions.

    PubMed

    Górski, A; Castronovo, V; Stepień-Sopniewska, B; Grieb, P; Ryba, M; Mrowiec, T; Korczak-Kowalska, G; Wierzbicki, P; Matysiak, W; Dybowska, B

    1994-07-01

    Although T cells infiltrate malignant tumors, the local immune response is usually inefficient and tumors escape destruction. While extracellular matrix proteins strongly costimulate T cell responses in normal individuals, our studies indicate that peripheral blood T cells from cancer patients and tumor infiltrating cells respond poorly or are resistant to stimulative signals mediated by collagen I and IV and fibronectin. Moreover, the adhesive properties of cancer T cells are markedly depressed. Those functional deficiencies are paralleled by variable deficits in integrin and non-integrin T cell receptors for extracellular matrix. Immunotherapy with BCG causes a dramatic but transient increase in T cell: ECM interactions. PMID:7827959

  7. Guided extracellular matrix formation from fibroblast cells cultured on bio-inspired configurable multiscale substrata

    PubMed Central

    Bae, Won-Gyu; Kim, Jangho; Choung, Yun-Hoon; Chung, Yesol; Suh, Kahp Y.; Pang, Changhyun; Chung, Jong Hoon; Jeong, Hoon Eui

    2015-01-01

    Engineering complex extracellular matrix (ECM) is an important challenge for cell and tissue engineering applications as well as for understanding fundamental cell biology. We developed the methodology for fabrication of precisely controllable multiscale hierarchical structures using capillary force lithography in combination with original wrinkling technique for the generation of well-defined native ECM-like platforms by culturing fibroblast cells on the multiscale substrata [1]. This paper provides information on detailed characteristics of polyethylene glycol-diacrylate multiscale substrata. In addition, a possible model for guided extracellular matrix formation from fibroblast cells cultured on bio-inspired configurable multiscale substrata is proposed. PMID:26543882

  8. Human mesenchymal stem cells seeded on extracellular matrix-scaffold: viability and osteogenic potential.

    PubMed

    Penolazzi, Letizia; Mazzitelli, Stefania; Vecchiatini, Renata; Torreggiani, Elena; Lambertini, Elisabetta; Johnson, Scott; Badylak, Stephen F; Piva, Roberta; Nastruzzi, Claudio

    2012-02-01

    The development and the optimization of novel culture systems of mesenchymal osteoprogenitors are some of the most important challenges in the field of bone tissue engineering (TE). A new combination between cells and extracellular matrix (ECM)-scaffold, containing ECM has here been analyzed. As source for osteoprogenitors, mesenchymal stem cells obtained from human umbilical cord Wharton's Jelly (hWJMSCs), were used. As ECM-scaffold, a powder form of isolated and purified porcine urinary bladder matrix (pUBM), was employed. The goals of the current work were: (1) the characterization of the in vitro hWJMSCs behavior, in terms of viability, proliferation, and adhesion to ECM-scaffold; (2) the effectiveness of ECM-scaffold to induce/modulate the osteoblastic differentiation; and (3) the proposal for a possible application of cells/ECM-scaffold construct to the field of cell/TE. In this respect, the properties of the pUBM-scaffold in promoting and guiding the in vitro adhesion, proliferation, and three-dimensional colonization of hWJMSCs, without altering viability and morphological characteristics of the cells, are here described. Finally, we have also demonstrated that pUBM-scaffolds positively affect the expression of typical osteoblastic markers in hWJMSCs. PMID:21830215

  9. Osteoblasts extracellular matrix induces vessel like structures through glycosylated collagen I

    SciTech Connect

    Palmieri, D.; Valli, M.; Viglio, S.; Ferrari, N.; Ledda, B.; Volta, C.; Manduca, P.

    2010-03-10

    Extracellular matrix (ECM) plays a fundamental role in angiogenesis affecting endothelial cells proliferation, migration and differentiation. Vessels-like network formation in vitro is a reliable test to study the inductive effects of ECM on angiogenesis. Here we utilized matrix deposed by osteoblasts as substrate where the molecular and structural complexity of the endogenous ECM is preserved, to test if it induces vessel-like network formation by endothelial cells in vitro. ECM is more similar to the physiological substrate in vivo than other substrates previously utilized for these studies in vitro. Osteogenic ECM, prepared in vitro from mature osteoblasts at the phase of maximal deposition and glycosylation of collagen I, induces EAhy926, HUVEC, and HDMEC endothelial cells to form vessels-like structures and promotes the activation of metalloproteinase-2 (MMP-2); the functionality of the p-38/MAPK signaling pathway is required. Osteogenic ECM also induces a transient increase of CXCL12 and a decrease of the receptor CXCR4. The induction of vessel-like networks is dependent from proper glycosylation of collagens and does not occur on osteogenic ECMs if deglycosylated by -galactosidase or on less glycosylated ECMs derived from preosteoblasts and normal fibroblasts, while is sustained on ECM from osteogenesis imperfecta fibroblasts only when their mutation is associated with over-glycosylation of collagen type I. These data support that post-translational glycosylation has a role in the induction in endothelial cells in vitro of molecules conductive to self-organization in vessels-like structures.

  10. Novel insights into the function and dynamics of extracellular matrix in liver fibrosis.

    PubMed

    Karsdal, Morten A; Manon-Jensen, Tina; Genovese, Federica; Kristensen, Jacob H; Nielsen, Mette J; Sand, Jannie Marie B; Hansen, Niels-Ulrik B; Bay-Jensen, Anne-Christine; Bager, Cecilie L; Krag, Aleksander; Blanchard, Andy; Krarup, Henrik; Leeming, Diana J; Schuppan, Detlef

    2015-05-15

    Emerging evidence suggests that altered components and posttranslational modifications of proteins in the extracellular matrix (ECM) may both initiate and drive disease progression. The ECM is a complex grid consisting of multiple proteins, most of which play a vital role in containing the essential information needed for maintenance of a sophisticated structure anchoring the cells and sustaining normal function of tissues. Therefore, the matrix itself may be considered as a paracrine/endocrine entity, with more complex functions than previously appreciated. The aims of this review are to 1) explore key structural and functional components of the ECM as exemplified by monogenetic disorders leading to severe pathologies, 2) discuss selected pathological posttranslational modifications of ECM proteins resulting in altered functional (signaling) properties from the original structural proteins, and 3) discuss how these findings support the novel concept that an increasing number of components of the ECM harbor signaling functions that can modulate fibrotic liver disease. The ECM entails functions in addition to anchoring cells and modulating their migratory behavior. Key ECM components and their posttranslational modifications often harbor multiple domains with different signaling potential, in particular when modified during inflammation or wound healing. This signaling by the ECM should be considered a paracrine/endocrine function, as it affects cell phenotype, function, fate, and finally tissue homeostasis. These properties should be exploited to establish novel biochemical markers and antifibrotic treatment strategies for liver fibrosis as well as other fibrotic diseases. PMID:25767261

  11. Expression of extracellular matrix molecules typical of articular cartilage in the human scapholunate interosseous ligament

    PubMed Central

    Milz, S; Aktas, T; Putz, R; Benjamin, M

    2006-01-01

    The scapholunate interosseous ligament (SLIL) connects the scaphoid and lunate bones and plays a crucial role in carpal kinematics. Its rupture leads to carpal instability and impairment of radiocarpal joint function. As the ligament is one of the first structures affected in rheumatoid arthritis, we conducted an immunohistochemical study of cadaveric tissue to determine whether it contains known autoantigens for rheumatoid arthritis. We immunolabelled the ligament from one hand in 12 cadavers with monoclonal antibodies directed against a wide range of extracellular matrix (ECM) molecules associated with both fibrous and cartilaginous tissues. The labelling profile has also enabled us to comment on how the molecular composition of the ligament relates to its mechanical function. All regions of the ligament labelled for types I, III and VI collagens, chondroitin 4 and 6 sulphates, keratan sulphate, dermatan sulphate, versican, tenascin and cartilage oligomeric matrix protein (COMP). However, both entheses labelled strongly for type II collagen, aggrecan and link protein and were distinctly fibrocartilaginous. In some regions, the ligament attached to bone via a region of hyaline cartilage that was continuous with articular cartilage. Labelling for cartilage molecules in the midsubstance was most evident dorsally. We conclude that the SLIL has an ECM which is typical of other highly fibrocartilaginous ligaments that experience both tensile load and shear. The presence of aggrecan, link protein, COMP and type II collagen could explain why the ligament may be a target for autoantigenic destruction in some forms of rheumatoid arthritis. PMID:16761970

  12. Novel insights into the function and dynamics of extracellular matrix in liver fibrosis

    PubMed Central

    Manon-Jensen, Tina; Genovese, Federica; Kristensen, Jacob H.; Nielsen, Mette J.; Sand, Jannie Marie B.; Hansen, Niels-Ulrik B.; Bay-Jensen, Anne-Christine; Bager, Cecilie L.; Krag, Aleksander; Blanchard, Andy; Krarup, Henrik; Leeming, Diana J.; Schuppan, Detlef

    2015-01-01

    Emerging evidence suggests that altered components and posttranslational modifications of proteins in the extracellular matrix (ECM) may both initiate and drive disease progression. The ECM is a complex grid consisting of multiple proteins, most of which play a vital role in containing the essential information needed for maintenance of a sophisticated structure anchoring the cells and sustaining normal function of tissues. Therefore, the matrix itself may be considered as a paracrine/endocrine entity, with more complex functions than previously appreciated. The aims of this review are to 1) explore key structural and functional components of the ECM as exemplified by monogenetic disorders leading to severe pathologies, 2) discuss selected pathological posttranslational modifications of ECM proteins resulting in altered functional (signaling) properties from the original structural proteins, and 3) discuss how these findings support the novel concept that an increasing number of components of the ECM harbor signaling functions that can modulate fibrotic liver disease. The ECM entails functions in addition to anchoring cells and modulating their migratory behavior. Key ECM components and their posttranslational modifications often harbor multiple domains with different signaling potential, in particular when modified during inflammation or wound healing. This signaling by the ECM should be considered a paracrine/endocrine function, as it affects cell phenotype, function, fate, and finally tissue homeostasis. These properties should be exploited to establish novel biochemical markers and antifibrotic treatment strategies for liver fibrosis as well as other fibrotic diseases. PMID:25767261

  13. Degenerated human intervertebral discs contain autoantibodies against extracellular matrix proteins.

    PubMed

    Capossela, S; Schläfli, P; Bertolo, A; Janner, T; Stadler, B M; Pötzel, T; Baur, M; Stoyanov, J V

    2014-01-01

    Degeneration of intervertebral discs (IVDs) is associated with back pain and elevated levels of inflammatory cells. It has been hypothesised that discogenic pain is a direct result of vascular and neural ingrowth along annulus fissures, which may expose the avascular nucleus pulposus (NP) to the systemic circulation and induce an autoimmune reaction. In this study, we confirmed our previous observation of antibodies in human degenerated and post-traumatic IVDs cultured in vitro. We hypothesised that the presence of antibodies was due to an autoimmune reaction against specific proteins of the disc. Furthermore we identified antigens which possibly trigger an autoimmune response in degenerative disc diseases. We demonstrated that degenerated and post-traumatic IVDs contain IgG antibodies against typical extracellular proteins of the disc, particularly proteins of the NP. We identified IgGs against collagen type II and aggrecan, confirming an autoimmune reaction against the normally immune privileged NP. We also found specific IgGs against collagens types I and V, but not against collagen type III. In conclusion, this study confirmed the association between disc degeneration and autoimmunity, and may open the avenue for future studies on developing prognostic, diagnostic and therapy-monitoring markers for degenerative disc diseases. PMID:24706108

  14. Alterations in the biosynthesis of extracellular matrix molecules in connective tissues by electric and magnetic fields

    SciTech Connect

    Ciombor, D.M.

    1992-01-01

    Pulsed electromagnetic fields (PEMFs) of certain configurations have been shown to be effective clinically in promoting the healing of fracture non-unions and are believed to enhance calcification of extracellular matrix. In vitro studies have suggested that PEMFs may also have the effect of modifying the extracellular matrix by promoting the synthesis of matrix molecules. This study examines the effect of one particular type of PEMF and a sinusoidal continuous wave upon the extracellular matrix and calcification of endochondral ossification in vivo. The pulsed magnetic field (SS-22) utilized in these studies is being used clinically for the treatment of fracture non-unions, a condition in which the bone is not restored to form or function. The sinusoidal continuous wave was designed to provide a 5 Gauss amplitude at a 15 Hz. rate. The synthesis of cartilage molecules is enhanced by this type of PEMF and since wave and subsequent endochondral calcification is stimulated. Histomorphometric studies indicate that the maturation of bone trabeculae is also promoted by this type of PEMF stimulation. These results indicate that a specific PEMF or continuous waveform can change the composition of cartilage extracellular matrix in vivo and raises the possibility that the effects on other processes of endochondral ossification (e.g., fracture healing and growth plates) may occur through a similar mechanism.

  15. Effect of insoluble extracellular matrix molecules on Fas expression in epithelial cells.

    PubMed

    Fine, A; Miranda, K; Farmer, S R; Anderson, N L

    1998-03-01

    Fas, which functions to initiate a signal causing apoptosis, is expressed in epithelia, thus, suggesting a role in controlling cell number during states of cell and matrix turnover. In view of this, we hypothesized that cell-matrix interactions may be an important determinant of Fas expression in epithelial cells. To investigate this, we examined the effect of insoluble extracellular matrix molecules on Fas expression in murine lung epithelial (MLE) cells, a transformed mouse lung epithelial cell line. We report that 1) insoluble extracellular matrices increased Fas mRNA in a time and concentration-dependent manner; 2) induced increases in Fas mRNA were associated with concomitantly increased Fas protein; and 3) nonspecific adherence to a polylysine substrate did not induce Fas mRNA. Consistent with these findings, Fas-induced apoptosis was significantly enhanced in cultures plated on type IV collagen. Employing rat hepatocytes, we confirmed that the insoluble extracellular matrix also increases Fas expression in primary epithelial cells. By amplifying Fas-mediated apoptosis, these data suggest a mechanism whereby the extracellular matrix regulates the fate of specific epithelial cell populations. PMID:9462690

  16. Printing three-dimensional tissue analogues with decellularized extracellular matrix bioink

    NASA Astrophysics Data System (ADS)

    Pati, Falguni; Jang, Jinah; Ha, Dong-Heon; Won Kim, Sung; Rhie, Jong-Won; Shim, Jin-Hyung; Kim, Deok-Ho; Cho, Dong-Woo

    2014-06-01

    The ability to print and pattern all the components that make up a tissue (cells and matrix materials) in three dimensions to generate structures similar to tissues is an exciting prospect of bioprinting. However, the majority of the matrix materials used so far for bioprinting cannot represent the complexity of natural extracellular matrix (ECM) and thus are unable to reconstitute the intrinsic cellular morphologies and functions. Here, we develop a method for the bioprinting of cell-laden constructs with novel decellularized extracellular matrix (dECM) bioink capable of providing an optimized microenvironment conducive to the growth of three-dimensional structured tissue. We show the versatility and flexibility of the developed bioprinting process using tissue-specific dECM bioinks, including adipose, cartilage and heart tissues, capable of providing crucial cues for cells engraftment, survival and long-term function. We achieve high cell viability and functionality of the printed dECM structures using our bioprinting method.

  17. Printing three-dimensional tissue analogues with decellularized extracellular matrix bioink

    PubMed Central

    Pati, Falguni; Jang, Jinah; Ha, Dong-Heon; Won Kim, Sung; Rhie, Jong-Won; Shim, Jin-Hyung; Kim, Deok-Ho; Cho, Dong-Woo

    2014-01-01

    The ability to print and pattern all the components that make up a tissue (cells and matrix materials) in three dimensions to generate structures similar to tissues is an exciting prospect of bioprinting. However, the majority of the matrix materials used so far for bioprinting cannot represent the complexity of natural extracellular matrix (ECM) and thus are unable to reconstitute the intrinsic cellular morphologies and functions. Here, we develop a method for the bioprinting of cell-laden constructs with novel decellularized extracellular matrix (dECM) bioink capable of providing an optimized microenvironment conducive to the growth of three-dimensional structured tissue. We show the versatility and flexibility of the developed bioprinting process using tissue-specific dECM bioinks, including adipose, cartilage and heart tissues, capable of providing crucial cues for cells engraftment, survival and long-term function. We achieve high cell viability and functionality of the printed dECM structures using our bioprinting method. PMID:24887553

  18. Extracellular matrix structure and nano-mechanics determine megakaryocyte function.

    PubMed

    Malara, Alessandro; Gruppi, Cristian; Pallotta, Isabella; Spedden, Elise; Tenni, Ruggero; Raspanti, Mario; Kaplan, David; Tira, Maria Enrica; Staii, Cristian; Balduini, Alessandra

    2011-10-20

    Cell interactions with matrices via specific receptors control many functions, with chemistry, physics, and membrane elasticity as fundamental elements of the processes involved. Little is known about how biochemical and biophysical processes integrate to generate force and, ultimately, to regulate hemopoiesis into the bone marrow-matrix environment. To address this hypothesis, in this work we focus on the regulation of MK development by type I collagen. By atomic force microscopy analysis, we demonstrate that the tensile strength of fibrils in type I collagen structure is a fundamental requirement to regulate cytoskeleton contractility of human MKs through the activation of integrin-α2β1-dependent Rho-ROCK pathway and MLC-2 phosphorylation. Most importantly, this mechanism seemed to mediate MK migration, fibronectin assembly, and platelet formation. On the contrary, a decrease in mechanical tension caused by N-acetylation of lysine side chains in type I collagen completely reverted these processes by preventing fibrillogenesis. PMID:21828129

  19. Effect of advanced glycation end products, extracellular matrix metalloproteinase inducer and matrix metalloproteinases on type-I collagen metabolism

    PubMed Central

    LI, WANG; LING, WANG; TENG, XIAOMEI; QUAN, CUIXIA; CAI, SHENGNAN; HU, SHUQUN

    2016-01-01

    The aim of the study was to examine the association among advanced glycation end products (AGEs), extracellular matrix metalloproteinase inducer (EMMPRIN) and matrix metalloproteinase (MMPs), and investigate whether AGEs affect type I collagen (COL-I) through EMMPRIN or MMPs. A co-culture system with the osteoblast-like cells (MC3T3E1) and mouse RAW264.7 cells was employed to examine the effects of AGE-bovine serum albumin (BSA) (50 mg/l), EMMPRIN antibody (5 mg/l) and AGE-BSA+EMMPRIN antibody separately on COL-I expression for 24 h. Culture media were analyzed for the content of COL-I by ELISA. The effect of different concentrations of AGE-BSA (0, 50, 100, 200 and 400 mg/l) for 24 h was assessed on COL-I levels. Finally, semiquantitative RT-PCR was used to detect the osteoblast COL-I mRNA expression and MMP-2 and MMP-9's PMAO were also measured in the culture medium. COL-I content in the culture medium decreased significantly following treatment with AGE-BSA (P<0.05). EMMPRIN antibody increased COL-I content (P<0.05). EMMPRIN antibody+AGE-BSA increased COL-I significantly (P<0.05). Different concentrations of AGE-BSA increased COL-I mRNA expression significantly compared with the control group (P<0.05), and were enhanced with increasing AGE-BSA concentration (P<0.05). Also MMP-2 and MMP-9 secretion increased significantly (P<0.05), with the increasing AGE-BSA concentration. In conclusion, an increase in AGE levels in vitro stimulates the secretion of EMMPRIN/MMPs, promotes the degradation of COL-I and reduces bone strength. PMID:27284408

  20. Characterization of the Vibrio cholerae Extracellular Matrix: A Top-Down Solid-State NMR Approach

    PubMed Central

    Reichhardt, Courtney; Fong, Jiunn C.N.; Yildiz, Fitnat; Cegelski, Lynette

    2015-01-01

    Bacterial biofilms are communities of bacterial cells surrounded by a self-secreted extracellular matrix. Biofilm formation by Vibrio cholerae, the human pathogen responsible for cholera, contributes to its environmental survival and infectivity. Important genetic and molecular requirements have been identified for V. cholerae biofilm formation, yet a compositional accounting of these parts in the intact biofilm or extracellular matrix has not been described. As insoluble and non-crystalline assemblies, determinations of biofilm composition pose a challenge to conventional biochemical and biophysical analysis. The V. cholerae extracellular matrix composition is particularly complex with several proteins, complex polysaccharides, and other biomolecules having been identified as matrix parts. We developed a new top-down solid-state NMR approach to spectroscopically assign and quantify the carbon pools of the intact V. cholerae extracellular matrix using 13C CPMAS and 13C{15N}, 15N{31P}, and 13C{31P}REDOR. General sugar, lipid, and amino acid pools were first profiled and then further annotated and quantified as specific carbon types, including carbonyls, amides, glycyl carbons, and anomerics. In addition, 15N profiling revealed a large amine pool relative to amide contributions, reflecting the prevalence of molecular modifications with free amine groups. Our top-down approach could be implemented immediately to examine the extracellular matrix from mutant strains that might alter polysaccharide production or lipid release beyond the cell surface; or to monitor changes that may accompany environmental variations and stressors such as altered nutrient composition, oxidative stress or antibiotics. More generally, our analysis has demonstrated that solid-state NMR is a valuable tool to characterize complex biofilm systems. PMID:24911407

  1. Macromolecularly crowded in vitro microenvironments accelerate the production of extracellular matrix-rich supramolecular assemblies

    PubMed Central

    Kumar, Pramod; Satyam, Abhigyan; Fan, Xingliang; Collin, Estelle; Rochev, Yury; Rodriguez, Brian J.; Gorelov, Alexander; Dillon, Simon; Joshi, Lokesh; Raghunath, Michael; Pandit, Abhay; Zeugolis, Dimitrios I.

    2015-01-01

    Therapeutic strategies based on the principles of tissue engineering by self-assembly put forward the notion that functional regeneration can be achieved by utilising the inherent capacity of cells to create highly sophisticated supramolecular assemblies. However, in dilute ex vivo microenvironments, prolonged culture time is required to develop an extracellular matrix-rich implantable device. Herein, we assessed the influence of macromolecular crowding, a biophysical phenomenon that regulates intra- and extra-cellular activities in multicellular organisms, in human corneal fibroblast culture. In the presence of macromolecules, abundant extracellular matrix deposition was evidenced as fast as 48 h in culture, even at low serum concentration. Temperature responsive copolymers allowed the detachment of dense and cohesive supramolecularly assembled living substitutes within 6 days in culture. Morphological, histological, gene and protein analysis assays demonstrated maintenance of tissue-specific function. Macromolecular crowding opens new avenues for a more rational design in engineering of clinically relevant tissue modules in vitro. PMID:25736020

  2. Development-dependent modification of the extracellular matrix by a sulphated glycoprotein in Volvox carteri

    PubMed Central

    Wenzl, Stephan; Thym, Detlef; Sumper, Manfred

    1984-01-01

    We report the chemical characterization of the highly sulphated glycoprotein SSG 185 from Volvox carteri. SSG 185 is a hydroxyproline-containing, extracellular glycoprotein. The sulphate residues are clustered within the parent saccharide structure of SSG 185, since on mercaptolysis all the sulphate residues are recovered in a small saccharide fragment containing mannose, arabinose and sulphate (in a molar ratio of 1̇1̇2). SSG 185 is a short-lived molecule, serving as a precursor for a high mol. wt. component of the extracellular matrix. Synthesis of SSG 185 is developmentally controlled. Different SSG 185 variants, with unknown modifications in the sulphated saccharide fragment, are synthesized at different developmental stages or under the influence of the sexual inducer. These modifications remain conserved in the aggregated state of SSG 185, indicating the development-dependent modification of the extracellular matrix. ImagesFig. 2.Fig. 3.Fig. 4.Fig. 6.Fig. 7. PMID:16453512

  3. Cyclopamine disrupts tumor extracellular matrix and improves the distribution and efficacy of nanotherapeutics in pancreatic cancer.

    PubMed

    Zhang, Bo; Jiang, Ting; Shen, Shun; She, Xiaojian; Tuo, Yanyan; Hu, Yu; Pang, Zhiqing; Jiang, Xinguo

    2016-10-01

    The dense extracellular matrix in pancreatic ductal adenocarcinoma dramatically reduces the penetration and efficacy of nanotherapeutics. Disruption of the tumor extracellular matrix may help improve the distribution and efficacy of nanotherapeutics in pancreatic cancer. In this study, we tested whether cyclopamine, a special inhibitor of the hedgehog signaling pathway with powerful anti-fibrotic activity, could promote the penetration and efficacy of nanotherapeutics in pancreatic cancer. It was shown that cyclopamine disrupted tumor extracellular fibronectins, decompressed tumor blood vessels, and improved tumor perfusion. Furthermore, cyclopamine improved the accumulation and intratumoral distribution of i.v.-administered fluorescence indicator-labeled nanoparticles. Finally, cyclopamine also significantly improved the tumor growth inhibition effect of i.v.-injected nanotherapeutics in pancreatic tumor xenograft mouse models. Thus, cyclopamine may have great potential to improve the therapeutic effects of nanomedicine in patients with pancreatic cancer. PMID:27376555

  4. Macromolecularly crowded in vitro microenvironments accelerate the production of extracellular matrix-rich supramolecular assemblies.

    PubMed

    Kumar, Pramod; Satyam, Abhigyan; Fan, Xingliang; Collin, Estelle; Rochev, Yury; Rodriguez, Brian J; Gorelov, Alexander; Dillon, Simon; Joshi, Lokesh; Raghunath, Michael; Pandit, Abhay; Zeugolis, Dimitrios I

    2015-01-01

    Therapeutic strategies based on the principles of tissue engineering by self-assembly put forward the notion that functional regeneration can be achieved by utilising the inherent capacity of cells to create highly sophisticated supramolecular assemblies. However, in dilute ex vivo microenvironments, prolonged culture time is required to develop an extracellular matrix-rich implantable device. Herein, we assessed the influence of macromolecular crowding, a biophysical phenomenon that regulates intra- and extra-cellular activities in multicellular organisms, in human corneal fibroblast culture. In the presence of macromolecules, abundant extracellular matrix deposition was evidenced as fast as 48 h in culture, even at low serum concentration. Temperature responsive copolymers allowed the detachment of dense and cohesive supramolecularly assembled living substitutes within 6 days in culture. Morphological, histological, gene and protein analysis assays demonstrated maintenance of tissue-specific function. Macromolecular crowding opens new avenues for a more rational design in engineering of clinically relevant tissue modules in vitro. PMID:25736020

  5. First demonstration of decorin, an extracellular matrix molecule, in bovine mammary tissue

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In the mammary gland, the extracellular matrix (ECM) is secreted by and surrounds cells located in both mammary parenchyma (PAR) and stroma. Decorin is an ECM proteoglycan with cell growth regulatory effects mediated by its ability to interact with growth factors or up-regulation of cyclin-dependent...

  6. Dynamic compressive behavior of human meniscus correlates with its extra-cellular matrix composition.

    PubMed

    Bursac, P; Arnoczky, S; York, A

    2009-01-01

    The menisci of the knee play a significant role in the complex biomechanics of the joint and are critically important in maintaining articular cartilage health. While a general form-function relationship has been identified for the structural orientation of the extra-cellular matrix of the meniscus, the role of individual biochemical components has yet to be fully explored. To determine if correlations exist between the dynamic and static compressive modulus of human menisci and their major extra-cellular matrix constituents (collagen, glycosoaminoglycan and water content), 12 lateral and 11 medial menisci from 13 adult donors were examined. The results showed that in dynamic compression at high loading frequencies (0.1-1 Hz) the menisci behave as a rubber-like elastic material while at lower frequencies (0.01-0.03 Hz) significant viscous dissipation occurs. While regional variations in compressive moduli and extra-cellular matrix composition were observed, the magnitude of both dynamic and static compressive moduli were found to be insensitive to collagen content (p>0.4). However, this magnitude was found to significantly increase with increasing glycosaminoglycan content (p<0.001) and significantly decrease with increasing water content (p<0.001). The results of this study identify significant relationships between the viscoelastic behavior of the meniscus and its extra-cellular matrix composition. PMID:19581729

  7. Decellularized skeletal muscle as an in vitro model for studying drug-extracellular matrix interactions.

    PubMed

    Wassenaar, Jean W; Boss, Gerry R; Christman, Karen L

    2015-09-01

    Several factors can affect drug absorption after intramuscular (IM) injection: drug solubility, drug transport across cell membranes, and drug metabolism at the injection site. We found that potential interactions between the drug and the extracellular matrix (ECM) at the injection site can also affect the rate of absorption post-injection. Using decellularized skeletal muscle, we developed a simple method to model drug absorption after IM injection, and showed that the nature of the drug-ECM interaction could be investigated by adding compounds that alter binding. We validated the model using the vitamin B12 analog cobinamide with different bound ligands. Cobinamide is being developed as an IM injectable treatment for cyanide poisoning, and we found that the in vitro binding data correlated with previously published in vivo drug absorption in animals. Commercially available ECM products, such as collagen and GelTrex, did not recapitulate drug binding behavior. While decellularized ECM has been widely studied in fields such as tissue engineering, this work establishes a novel use of skeletal muscle ECM as a potential in vitro model to study drug-ECM interactions during drug development. PMID:26125502

  8. The neuronal extracellular matrix restricts distribution and internalization of aggregated Tau-protein.

    PubMed

    Suttkus, A; Holzer, M; Morawski, M; Arendt, T

    2016-01-28

    Alzheimer's disease (AD) is a chronic degenerative disorder characterized by fibrillary aggregates of Aß and Tau-protein. Formation and progression of these pathological hallmarks throughout the brain follow a specific spatio-temporal pattern which provides the basis for neuropathological staging. Previously, we could demonstrate that cortical and subcortical neurons are less frequently affected by neurofibrillary degeneration if they are enwrapped by a specialized form of the hyaluronan-based extracellular matrix (ECM), the so called 'perineuronal net' (PN). PNs are composed of large aggregating chondroitin sulfate proteoglycans connected to a hyaluronan backbone, stabilized by link proteins and cross-linked via tenascin-R. Recently, PN-associated neurons were shown to be better protected against iron-induced neurodegeneration compared to neurons without PN, indicating a neuroprotective function. Here, we investigated the role of PNs in distribution and internalization of exogenous Tau-protein by using organotypic slice cultures of wildtype mice as well as mice lacking the ECM-components aggrecan, HAPLN1 or tenascin-R. We could demonstrate that PNs restrict both distribution and internalization of Tau. Accordingly, PN-ensheathed neurons were less frequently affected by Tau-internalization, than neurons without PN. Finally, the PNs as well as their three investigated components were shown to modulate the processes of distribution as well as internalization of Tau. PMID:26621125

  9. Spatial organization of the extracellular matrix regulates cell–cell junction positioning

    PubMed Central

    Tseng, Qingzong; Duchemin-Pelletier, Eve; Deshiere, Alexandre; Balland, Martial; Guillou, Hervé; Filhol, Odile; Théry, Manuel

    2012-01-01

    The organization of cells into epithelium depends on cell interaction with both the extracellular matrix (ECM) and adjacent cells. The role of cell–cell adhesion in the regulation of epithelial topology is well-described. ECM is better known to promote cell migration and provide a structural scaffold for cell anchoring, but its contribution to multicellular morphogenesis is less well-understood. We developed a minimal model system to investigate how ECM affects the spatial organization of intercellular junctions. Fibronectin micropatterns were used to constrain the location of cell–ECM adhesion. We found that ECM affects the degree of stability of intercellular junction positioning and the magnitude of intra- and intercellular forces. Intercellular junctions were permanently displaced, and experienced large perpendicular tensional forces as long as they were positioned close to ECM. They remained stable solely in regions deprived of ECM, where they were submitted to lower tensional forces. The heterogeneity of the spatial organization of ECM induced anisotropic distribution of mechanical constraints in cells, which seemed to adapt their position to minimize both intra- and intercellular forces. These results uncover a morphogenetic role for ECM in the mechanical regulation of cells and intercellular junction positioning. PMID:22307605

  10. Physicomechanical properties of the extracellular matrix of a demineralized bone

    NASA Astrophysics Data System (ADS)

    Kirilova, I. A.; Sharkeev, Yu. P.; Nikolaev, S. V.; Podorozhnaya, V. T.; Uvarkin, P. V.; Ratushnyak, A. S.; Chebodaeva, V. V.

    2016-08-01

    The article describes the results of a study of physicomechanical properties of a demineralized bone matrix of human cancellous and compact bones. A demineralized cancellous bone was shown to have the best characteristics of a porous system for colonization of matrices by cells. The ultimate stress and elasticity modulus of samples of demineralized femoral heads isolated in primary hip replacement was demonstrated to vary in wide ranges. The elasticity modulus ranged from 50 to 250 MPa, and the tensile strength varied from 1.1 to 5.5 MPa. Microhardness measurements by the recovered indentation method were not possible because of the viscoelastic properties of a bone material. To study the piezoelectric properties of samples, a measuring system was developed that comprised a measuring chamber with contact electrodes, a system for controlled sample loading, an amplifier-converter unit, and signal recording and processing software. The measurement results were used to determine the dependence of the signal amplitude on the dynamic deformation characteristics. The findings are discussed in terms of the relationship between the mechanical and electrical properties and the structure of the organic bone component.

  11. Extracellular Matrix Molecular Remodeling in Human Liver Fibrosis Evolution

    PubMed Central

    Baiocchini, Andrea; Montaldo, Claudia; Conigliaro, Alice; Grimaldi, Alessio; Correani, Virginia; Mura, Francesco; Ciccosanti, Fabiola; Rotiroti, Nicolina; Brenna, Alessia; Montalbano, Marzia; D’Offizi, Gianpiero; Capobianchi, Maria Rosaria; Alessandro, Riccardo; Piacentini, Mauro; Schininà, Maria Eugenia; Maras, Bruno; Del Nonno, Franca; Tripodi, Marco; Mancone, Carmine

    2016-01-01

    Chronic liver damage leads to pathological accumulation of ECM proteins (liver fibrosis). Comprehensive characterization of the human ECM molecular composition is essential for gaining insights into the mechanisms of liver disease. To date, studies of ECM remodeling in human liver diseases have been hampered by the unavailability of purified ECM. Here, we developed a decellularization method to purify ECM scaffolds from human liver tissues. Histological and electron microscopy analyses demonstrated that the ECM scaffolds, devoid of plasma and cellular components, preserved the three-dimensional ECM structure and zonal distribution of ECM components. This method has been then applied on 57 liver biopsies of HCV-infected patients at different stages of liver fibrosis according to METAVIR classification. Label-free nLC-MS/MS proteomics and computation biology were performed to analyze the ECM molecular composition in liver fibrosis progression, thus unveiling protein expression signatures specific for the HCV-related liver fibrotic stages. In particular, the ECM molecular composition of liver fibrosis was found to involve dynamic changes in matrix stiffness, flexibility and density related to the dysregulation of predominant collagen, elastic fibers and minor components with both structural and signaling properties. This study contributes to the understanding of the molecular bases underlying ECM remodeling in liver fibrosis and suggests new molecular targets for fibrolytic strategies. PMID:26998606

  12. Neoplastic extracellular matrix environment promotes cancer invasion in vitro.

    PubMed

    Sundquist, Elias; Renko, Outi; Salo, Sirpa; Magga, Johanna; Cervigne, Nilva K; Nyberg, Pia; Risteli, Juha; Sormunen, Raija; Vuolteenaho, Olli; Zandonadi, Flávia; Paes Leme, Adriana F; Coletta, Ricardo D; Ruskoaho, Heikki; Salo, Tuula

    2016-06-10

    The invasion of carcinoma cells is a crucial feature in carcinogenesis. The penetration efficiency not only depends on the cancer cells, but also on the composition of the tumor microenvironment. Our group has developed a 3D invasion assay based on human uterine leiomyoma tissue. Here we tested whether human, porcine, mouse or rat hearts as well as porcine tongue tissues could be similarly used to study carcinoma cell invasion in vitro. Three invasive human oral tongue squamous cell carcinoma (HSC-3, SCC-25 and SCC-15), melanoma (G-361) and ductal breast adenocarcinoma (MDA-MB-231) cell lines, and co-cultures of HSC-3 and carcinoma-associated or normal oral fibroblasts were assayed. Myoma tissue, both native and lyophilized, promoted invasion and growth of the cancer cells. However, the healthy heart or tongue matrices were unable to induce the invasion of any type of cancer cells tested. Moreover, when studied in more detail, small molecular weight fragments derived from heart tissue rinsing media inhibited HSC-3 horizontal migration. Proteome analysis of myoma rinsing media, on the other hand, revealed migration enhancing factors. These results highlight the important role of matrix composition for cancer invasion studies in vitro and further demonstrate the unique properties of human myoma organotypic model. PMID:27090016

  13. Inhaled Steroids Modulate Extracellular Matrix Composition in Bronchial Biopsies of COPD Patients: A Randomized, Controlled Trial

    PubMed Central

    Kunz, Lisette I. Z.; Strebus, Jolanda; Budulac, Simona E.; Lapperre, Therese S.; Sterk, Peter J.; Postma, Dirkje S.; Mauad, Thais; Timens, Wim; Hiemstra, Pieter S.

    2013-01-01

    Rationale Smoking and inflammation contribute to the pathogenesis of chronic obstructive pulmonary disease (COPD), which involves changes in extracellular matrix. This is thought to contribute to airway remodeling and airflow obstruction. We have previously observed that long-term treatment with inhaled corticosteroids can not only reduce bronchial inflammation, but can also attenuate lung function decline in moderate-severe COPD. We hypothesized that inhaled corticosteroids and current smoking modulate bronchial extracellular matrix components in COPD. Objective To compare major extracellular matrix components (elastic fibers; proteoglycans [versican, decorin]; collagens type I and III) in bronchial biopsies 1) after 30-months inhaled steroids treatment or placebo; and 2) between current and ex-smokers with COPD. Methods We included 64 moderate-severe, steroid-naive COPD patients (24/40 (ex)-smokers, 62±7 years, 46 (31–54) packyears, post-bronchodilator forced expiratory volume in one second (FEV1) 62±9% predicted) at baseline in this randomized, controlled trial. 19 and 13 patients received 30-months treatment with fluticasone or placebo, respectively. Bronchial biopsies collected at baseline and after 30 months were studied using (immuno)histochemistry to evaluate extracellular matrix content. Percentage and density of stained area were calculated by digital image analysis. Results 30-Months inhaled steroids increased the percentage stained area of versican (9.6% [CI 0.9 to 18.3%]; p = 0.03) and collagen III (20.6% [CI 3.8 to 37.4%]; p = 0.02) compared to placebo. Increased collagen I staining density correlated with increased post-bronchodilator FEV1 after inhaled steroids treatment (Rs = 0.45, p = 0.04). There were no differences between smokers and ex-smokers with COPD in percentages and densities for all extracellular matrix proteins. Conclusions These data show that long-term inhaled corticosteroids treatment partially changes the

  14. Local fluid transfer regulation in heart extracellular matrix.

    PubMed

    McGee, Maria P; Morykwas, Michael J; Jordan, James E; Wang, Rui; Argenta, Louis C

    2016-06-01

    The interstitial myocardial matrix is a complex and dynamic structure that adapts to local fluctuations in pressure and actively contributes to the heart's fluid exchange and hydration. However, classical physiologic models tend to treat it as a passive conduit for water and solute, perhaps because local interstitial regulatory mechanisms are not easily accessible to experiment in vivo. Here, we examined the interstitial contribution to the fluid-driving pressure ex vivo. Interstitial hydration potentials were determined from influx/efflux rates measured in explants from healthy and ischemia-reperfusion-injured pigs during colloid osmotic pressure titrations. Adaptive responses were further explored by isolating myocardial fibroblasts and measuring their contractile responses to water activity changes in vitro. Results show hydration potentials between 5 and 60 mmHg in healthy myocardia and shifts in excess of 200 mmHg in edematous myocardia after ischemia-reperfusion injury. Further, rates of fluid transfer were temperature-dependent, and in collagen gel contraction assays, myocardial fibroblasts tended to preserve the micro-environment's hydration volume by slowing fluid efflux rates at pressures above 40 mmHg. Our studies quantify components of the fluid-driving forces in the heart interstitium that the classical Starling's equation does not explicitly consider. Measured hydration potentials in healthy myocardia and shifts with edema are larger than predicted from the known values of hydrostatic and colloid osmotic interstitial fluid pressures. Together with fibroblast responses in vitro, they are consistent with regulatory mechanisms that add local biological controls to classic fluid-balance models. PMID:26961911

  15. Electric field stimulation through a substrate influences Schwann cell and extracellular matrix structure

    NASA Astrophysics Data System (ADS)

    Nguyen, Hieu T.; Wei, Claudia; Chow, Jacqueline K.; Nguy, Lindsey; Nguyen, Hieu K.; Schmidt, Christine E.

    2013-08-01

    Objective. Electric field (EF) stimulation has been used to cue cell growth for tissue engineering applications. In this study, we explore the electrical parameters and extracellular mechanisms that elicit changes in cell behavior when stimulated through the substrate. Approach. Rat Schwann cell morphology was compared when exposed to EF through the media or a conductive indium tin oxide substrate. Ionic and structural effects were then analyzed on Matrigel and collagen I, respectively. Main results. When stimulating through media, cells had greater alignment perpendicular to the EF with higher current densities (106 mA cm-2 at 245 mV mm-1), and reached maximum alignment within 8 h. Stimulation through the substrate with EF (up to 110 mV mm-1) did not affect Schwann cell orientation, however the EF caused extracellular matrix (ECM) coatings on substrates to peel away, suggesting EF can physically change the ECM. Applying alternating current (ac) 2-1000 Hz signals through the media or substrate both caused cells to flatten and protrude many processes, without preferential alignment. Matrigel exposed to a substrate EF of 10 mV mm-1 for 2 h had a greater calcium concentration near the cathode, but quickly dissipated when the EF was removed. Schwann cells seeded 7 d after gels were exposed to substrate EF still aligned perpendicular to the EF direction. Microscopy of collagen I exposed to substrate EF shows alignment and bundling of fibrils. Significance. These findings demonstrate EF exposure can control Schwann cell alignment and morphology, change ECM bulk/surface architecture, and align ECM structures.

  16. Endometrial gene expression of acute phase extracellular matrix components following estrogen disruption of pregnancy in pigs.

    PubMed

    Ashworth, Morgan D; Ross, Jason W; Stein, Daniel; White, Frank; Geisert, Rodney D

    2010-12-01

    In pigs, administration of estrogen to gilts on Days 9 and 10 of pregnancy causes conceptus fragmentation and death between Days 15 and 18 of gestation. Conceptus degeneration is associated with breakdown of the microvilli surface glycocalyx on the lumenal epithelium (LE). We previously identified endometrial expression of inter-α-trypsin inhibitor (ITI) and hyaluronic acid (HA), which are key components of extracellular matrix (ECM), during the period of conceptus attachment to the uterine surface in the pig. Tumor necrosis factor-α-inducible protein-6 (TNFAIP6) serves as a linker for ECM expansion and is stimulated by prostaglandin E (PGE). We hypothesized that early estrogen administration alters the normal ECM components forming glycocalyx on the LE. Bred gilts (4 gilts/trt/day) were treated with either 5mg estradiol cypionate (E) or corn oil (CO) on Days 9 and 10 of gestation. The uterus was surgically removed on either Days 10, 12, 13, 15 and 17 of gestation and endometrial tissue snap frozen in liquid nitrogen. Endometrial tumor necrosis factor-α (TNF), TNFAIP6, interleukin 6 (IL6), and inter-α-trypsin inhibitor heavy chains (ITIH) were detected during early pregnancy thereby indicating all components for maintenance of the extracellular glycocalyx are present in the endometrium of pigs. However, only gene expression of ITIH2 was suppressed by E-treatment. TNFAIP6 protein was detected across all days of gestation but was not affected by E-treatment. The present study demonstrates that while the pig endometrium expresses key components of ECM only ITIH2 gene expression was altered by E-treatment. A decrease in ITIH2 could lead to the possible loss of the uterine glycocalyx leading to conceptus degeneration; however, other factors may be involved with the loss of glycocalyx during implantation in the pig following E-treatment. PMID:20850941

  17. Cigarette smoke enhances proliferation and extracellular matrix deposition by human fetal airway smooth muscle

    PubMed Central

    Vogel, Elizabeth R.; VanOosten, Sarah K.; Holman, Michelle A.; Hohbein, Danielle D.; Thompson, Michael A.; Vassallo, Robert; Pandya, Hitesh C.; Prakash, Y. S.

    2014-01-01

    Cigarette smoke is a common environmental insult associated with increased risk of developing airway diseases such as wheezing and asthma in neonates and children. In adults, asthma involves airway remodeling characterized by increased airway smooth muscle (ASM) cell proliferation and increased extracellular matrix (ECM) deposition, as well as airway hyperreactivity. The effects of cigarette smoke on remodeling and contractility in the developing airway are not well-elucidated. In this study, we used canalicular-stage (18–20 wk gestational age) human fetal airway smooth muscle (fASM) cells as an in vitro model of the immature airway. fASM cells were exposed to cigarette smoke extract (CSE; 0.5–1.5% for 24–72 h), and cell proliferation, ECM deposition, and intracellular calcium ([Ca2+]i) responses to agonist (histamine 10 μM) were used to evaluate effects on remodeling and hyperreactivity. CSE significantly increased cell proliferation and deposition of ECM molecules collagen I, collagen III, and fibronectin. In contrast, [Ca2+]i responses were not significantly affected by CSE. Analysis of key signaling pathways demonstrated significant increase in extracellular signal-related kinase (ERK) and p38 activation with CSE. Inhibition of ERK or p38 signaling prevented CSE-mediated changes in proliferation, whereas only ERK inhibition attenuated the CSE-mediated increase in ECM deposition. Overall, these results demonstrate that cigarette smoke may enhance remodeling in developing human ASM through hyperplasia and ECM production, thus contributing to development of neonatal and pediatric airway disease. PMID:25344066

  18. Morphological Characterization of Organized Extracellular Matrix Deposition by Ascorbic Acid-Stimulated Human Corneal Fibroblasts

    PubMed Central

    Guo, Xiaoqing; Hutcheon, Audrey E. K.; Melotti, Suzanna A.; Zieske, James D.; Trinkaus-Randall, Vickery; Ruberti, Jeffrey W.

    2016-01-01

    Purpose To characterize the structure and morphology of extracellular matrix (ECM) synthesized by untransformed, cultured human corneal fibroblasts in long-term cultures. Methods Human corneal stromal keratocytes were expanded in transwell culture in the presence of fetal bovine serum and a stable derivative of Vitamin C. The cells were allowed to synthesize a fibrillar ECM for up to five weeks. Constructs were assessed via light (phase contrast and differential interference contrast) and transmission (standard and quick freeze/deep etch) microscopy. Results Electron micrographs revealed stratified constructs with multiple parallel layers of cells and an extracellular matrix comprising parallel arrays of small, polydisperse fibrils (27–51 nm) which often alternate in direction. Differential interference contrast images demonstrated oriented ECM fibril arrays parallel to the plane of the construct while quick-freeze deep etch micrographs showed the details of the matrix interaction with fibroblasts via arrays of membrane surface structures. Conclusions Human keratocytes, cultured in a stable Vitamin C derivative, are capable of assembling extracellular matrix which comprise parallel arrays of ECM fibrils. The resulting constructs, which are highly cellular, exhibit morphology similar to the developing mammalian stroma where organized matrix is derived. The appearance of arrays of structures on the cell membranes suggest a role in the local organization of synthesized ECM. This model could provide critical insight into the fundamental processes which govern the genesis of organized connective tissues such as the cornea and may provide a scaffolding suitable for tissue-engineering a biomimetic stroma. PMID:17724187

  19. All-trans retinoic acid and extracellular Ca2+ differentially influence extracellular matrix production by human skin in organ culture.

    PubMed Central

    Varani, J.; Larson, B. K.; Perone, P.; Inman, D. R.; Fligiel, S. E.; Voorhees, J. J.

    1993-01-01

    Two-mm full-thickness punch biopsies of human skin were placed in organ culture in a serum-free, growth factor-free basal medium. Under conditions of low extracellular Ca2+ (0.15 mmol/L), the tissue quickly degenerated. However, degeneration was prevented when the extracellular Ca2+ concentration was increased to 1.4 mmol/L. The tissue remained histologically normal in appearance and biochemically active for up to 12 days. The addition of 3 mumol/L all-trans retinoic acid (RA) to the low-Ca2+ culture medium also prevented tissue degeneration. However, in contrast to what was seen in the presence of 1.4 mmol/L Ca2+, epidermal differentiation did not occur normally in the presence of RA. Rather, the upper layers of the epidermis routinely separated from the underlying basal cells. Fibronectin production by the organ cultured skin was examined. Biosynthetic labeling/immunoprecipitation studies demonstrated that incubation of the tissue in basal medium containing 1.4 mmol/L Ca2+ resulted in a high level of fibronectin production relative to the amount produced in basal medium containing 0.15 mmol/L Ca2+. In contrast, the addition of 3 mumol/L RA to the low Ca2+ basal medium did not stimulate fibronectin production. Similar results were observed in enzyme-linked immunosorbent assays where the addition of Ca2+ to a final concentration of 1.4 mmol/L stimulated fibronectin and thrombospondin production whereas RA (3 mumol/L) did not. Although RA by itself failed to stimulate extracellular matrix production, the addition of 3 mumol/L RA to basal medium containing 1.4 mmol/L Ca2+ led to a further increase in fibronectin production over that seen in the presence of 1.4 mmol/L Ca2+ alone. Taken together, these data indicate that although either 1.4 mmol/L Ca2+ or 3 mumol/L RA facilitates survival of organ-cultured skin in basal medium, they have very different effects on extracellular matrix production. This supports the view, based on histological appearance, that the two

  20. Decellularized Tracheal Extracellular Matrix Supports Epithelial Migration, Differentiation, and Function

    PubMed Central

    Kutten, Johannes C.; McGovern, David; Hobson, Christopher M.; Luffy, Sarah A.; Nieponice, Alejandro; Tobita, Kimimasa; Francis, Richard J.; Reynolds, Susan D.

    2015-01-01

    Tracheal loss is a source of significant morbidity for affected patients with no acceptable solution. Interest in engineering tracheal transplants has created a demand for small animal models of orthotopic tracheal transplantation. Here, we examine the use of a decellularized graft in a murine model of tracheal replacement. Fresh or decellularized tracheas harvested from age-matched female donor C57BL/6 mice were transplanted into syngeneic recipients. Tracheas were decellularized using repeated washes of water, 3% Triton X-100, and 3 M NaCl under cyclic pressure changes, followed by disinfection with 0.1% peracetic acid/4% ethanol, and terminal sterilization by gamma irradiation. Tracheas were explanted for immunolabeling at 1, 4, and 8 weeks following surgery. Video microscopy and computed tomography were performed to assess function and structure. Decellularized grafts supported complete reepithelialization by 8 weeks and motile cilia were observed. Cartilaginous portions of the trachea were maintained in mice receiving fresh transplants, but repopulation of the cartilage was not seen in mice receiving decellularized transplants. We observed superior postsurgical survival, weight gain, and ciliary function in mice receiving fresh transplants compared with those receiving decellularized transplants. The murine orthotopic tracheal transplant provides an appropriate model to assess the repopulation and functional regeneration of decellularized tracheal grafts. PMID:24980864

  1. The extracellular-matrix-retaining cyanobacterium Nostoc verrucosum accumulates trehalose, but is sensitive to desiccation.

    PubMed

    Sakamoto, Toshio; Kumihashi, Keisuke; Kunita, Shinpei; Masaura, Takuya; Inoue-Sakamoto, Kaori; Yamaguchi, Masaaki

    2011-08-01

    The aquatic cyanobacterium Nostoc verrucosum forms macroscopic colonies, which consist of both cellular filaments and massive extracellular matrix material. In this study, the physiological features of N. verrucosum were investigated and compared with those of the anhydrobiotic cyanobacterium Nostoc commune. Nostoc verrucosum cells were sensitive to desiccation, but tolerant of freeze-thawing treatment in terms of both cell viability and photosynthetic O(2) evolution. Natural colonies of these cyanobacteria contained similar levels of chlorophyll a, carotenoids, the UV-absorbing pigments scytonemin and mycosporine-like amino acids, and uronic acid [a component of extracellular polysaccharides (EPS)]. EPS from both N. verrucosum and N. commune indicated low acidity and a high affinity for divalent cations, although their sugar compositions differed. The WspA protein, known to be a major component of the extracellular matrix of N. commune, was detected in N. verrucosum. Desiccation caused similarly high levels of trehalose accumulation in both cyanobacteria. Although previously considered relevant to anhydrobiosis in the terrestrial cyanobacterium N. commune, the data presented here suggest that extracellular matrix production and trehalose accumulation are not enough for standing extreme desiccation in N. verrucosum. PMID:21507024

  2. Changes in the expression of extracellular matrix (ECM) and matrix metalloproteinases (MMP) of proliferating rat parotid acinar cells.

    PubMed

    Broverman, R L; Nguyen, K H; da Silveira, A; Brinkley, L L; Macauley, S P; Zeng, T; Yamamoto, H; Tarnuzzer, R W; Schultz, G S; Kerr, M; Humphreys-Beher, M G

    1998-07-01

    Tissue morphogenesis, development, and maintenance of function are mediated by signals generated through the composition of the extracellular matrix. The regulation of the composition of matrix is determined by enzymes specific for their degradation, the matrix metalloproteinases. Chronic injections of the beta-adrenergic receptor agonist, isoproterenol, result in a non-neoplastic hypertrophy and hyperplasia of the rat parotid gland. The activity of matrix metalloproteinases, as measured by gelatin zymography and enzymatic digestion of Azocoll substrates by gland lysates, decreased significantly (P < 0.05) following 24 hrs of agonist treatment, and slowly recovered to control values by 6 days of treatment. Daily administration of the broad-spectrum matrix metalloproteinase inhibitor Galardin for 3 days in combination with isoproterenol resulted in enhanced gland hypertrophy compared with that produced by isoproterenol alone. Given alone, Galardin also caused hypertrophy. The relative abundance of mRNA for the extracellular matrix molecules, collagens I and III and fibronectin, declined rapidly following the initiation of beta-agonist treatment in vivo, while laminin B1 and B2 mRNA levels increased initially before declining below control levels. These changes in patterns of mRNA levels also were observed in the concentrations of glandular protein when Western dot blot analysis of collagens I and III and laminin, respectively, was used. The importance of laminin, in vivo, was demonstrated by coinjection of anti-laminin antibody along with isoproterenol, which resulted in the inhibition of beta-agonist-induced parotid gland hypertrophy and hyperplasia. These data suggest that modulation of the ECM is associated with isoproterenol-induced salivary gland hypertrophy and hyperplasia. It is likely that this modulation of the ECM takes place through transcriptional regulation of some ECM genes and regulation of matrix-degrading enzyme activity. PMID:9663435

  3. Cell response to RGD density in cross-linked artificial extracellular matrix protein films.

    PubMed

    Liu, Julie C; Tirrell, David A

    2008-11-01

    This study examines the adhesion, spreading, and migration of human umbilical vein endothelial cells on cross-linked films of artificial extracellular matrix (aECM) proteins. The aECM proteins described here were designed for application in small-diameter grafts and are composed of elastin-like structural repeats and fibronectin cell-binding domains. aECM-RGD contains the RGD sequence derived from fibronectin; the negative control protein aECM-RDG contains a scrambled cell-binding domain. The covalent attachment of poly(ethylene glycol) (PEG) to aECM substrates reduced nonspecific cell adhesion to aECM-RDG-PEG but did not preclude sequence-specific adhesion of endothelial cells to aECM-RGD-PEG. Variation in ligand density was accomplished by the mixing of aECM-RGD-PEG and aECM-RDG-PEG prior to cross-linking. Increasing the density of RGD domains in cross-linked films resulted in more robust cell adhesion and spreading but did not affect cell migration speed. Control of cell-binding domain density in aECM proteins can thus be used to modulate cell adhesion and spreading and will serve as an important design tool as these materials are further developed for use in surgery, tissue engineering, and regenerative medicine. PMID:18826275

  4. Fibroblasts behavior after N-acetylcysteine and amino acids exposure: extracellular matrix gene expression.

    PubMed

    Avantaggiato, Anna; Palmieri, Annalisa; Bertuzzi, Gianluigi; Carinci, Francesco

    2014-06-01

    Reactive oxygen species (ROS) are chemically reactive molecules with impaired electrons that make them unstable and able to react easily with a great variety of molecules. The main targets of ROS are DNA, proteins, and membrane phospholipids. In the skin, ROS are able to affect the production of collagen and elastin, the main components of the extracellular matrix (ECM). This action contributes to the skin's aging. N-Acetylcysteine (NAC) is an acetylated cysteine residue with excellent anti-oxidant activity that boosts glutathione (GSH) levels. This study evaluates the effect of a solution of NAC and amino acids, which is used in aesthetic medicine as an intra-dermal injective treatment, on fibroblast behavior. To this aim, the expression levels of some ECM-related genes (HAS1, HYAL1 ELN, ELANE, MMP2, MMP3, MMP13, COL1A1, COL3A1) were analyzed on cultured dermal fibroblasts using real-time reverse transcription polymerase chain reaction (RT-PCR). All but two collagen genes were up-regulated after 24 hr of treatment. PMID:24438160

  5. Vinculin is required for cell polarization, migration, and extracellular matrix remodeling in 3D collagen.

    PubMed

    Thievessen, Ingo; Fakhri, Nikta; Steinwachs, Julian; Kraus, Viola; McIsaac, R Scott; Gao, Liang; Chen, Bi-Chang; Baird, Michelle A; Davidson, Michael W; Betzig, Eric; Oldenbourg, Rudolf; Waterman, Clare M; Fabry, Ben

    2015-11-01

    Vinculin is filamentous (F)-actin-binding protein enriched in integrin-based adhesions to the extracellular matrix (ECM). Whereas studies in 2-dimensional (2D) tissue culture models have suggested that vinculin negatively regulates cell migration by promoting cytoskeleton-ECM coupling to strengthen and stabilize adhesions, its role in regulating cell migration in more physiologic, 3-dimensional (3D) environments is unclear. To address the role of vinculin in 3D cell migration, we analyzed the morphodynamics, migration, and ECM remodeling of primary murine embryonic fibroblasts (MEFs) with cre/loxP-mediated vinculin gene disruption in 3D collagen I cultures. We found that vinculin promoted 3D cell migration by increasing directional persistence. Vinculin was necessary for persistent cell protrusion, cell elongation, and stable cell orientation in 3D collagen, but was dispensable for lamellipodia formation, suggesting that vinculin-mediated cell adhesion to the ECM is needed to convert actin-based cell protrusion into persistent cell shape change and migration. Consistent with this finding, vinculin was necessary for efficient traction force generation in 3D collagen without affecting myosin II activity and promoted 3D collagen fiber alignment and macroscopical gel contraction. Our results suggest that vinculin promotes directionally persistent cell migration and tension-dependent ECM remodeling in complex 3D environments by increasing cell-ECM adhesion and traction force generation. PMID:26195589

  6. Extracellular matrix proteins expression profiling in chemoresistant variants of the A2780 ovarian cancer cell line.

    PubMed

    Januchowski, Radosław; Zawierucha, Piotr; Ruciński, Marcin; Nowicki, Michał; Zabel, Maciej

    2014-01-01

    Ovarian cancer is the leading cause of death among gynaecological malignancies. Extracellular matrix (ECM) can affect drug resistance by preventing the penetration of the drug into cancer cells and increased resistance to apoptosis. This study demonstrates alterations in the expression levels of ECM components and related genes in cisplatin-, doxorubicin-, topotecan-, and paclitaxel-resistant variants of the A2780 ovarian cancer cell line. Affymetrix Gene Chip Human Genome Array Strips were used for hybridisations. The genes that had altered expression levels in drug-resistant sublines were selected and filtered by scatter plots. The genes that were up- or downregulated more than fivefold were selected and listed. Among the investigated genes, 28 genes were upregulated, 10 genes were downregulated, and two genes were down- or upregulated depending on the cell line. Between upregulated genes 12 were upregulated very significantly--over 20-fold. These genes included COL1A2, COL12A1, COL21A1, LOX, TGFBI, LAMB1, EFEMP1, GPC3, SDC2, MGP, MMP3, and TIMP3. Four genes were very significantly downregulated: COL11A1, LAMA2, GPC6, and LUM. The expression profiles of investigated genes provide a preliminary insight into the relationship between drug resistance and the expression of ECM components. Identifying correlations between investigated genes and drug resistance will require further analysis. PMID:24804215

  7. Extracellular Matrix Proteins Expression Profiling in Chemoresistant Variants of the A2780 Ovarian Cancer Cell Line

    PubMed Central

    Januchowski, Radosław; Zawierucha, Piotr; Ruciński, Marcin; Nowicki, Michał; Zabel, Maciej

    2014-01-01

    Ovarian cancer is the leading cause of death among gynaecological malignancies. Extracellular matrix (ECM) can affect drug resistance by preventing the penetration of the drug into cancer cells and increased resistance to apoptosis. This study demonstrates alterations in the expression levels of ECM components and related genes in cisplatin-, doxorubicin-, topotecan-, and paclitaxel-resistant variants of the A2780 ovarian cancer cell line. Affymetrix Gene Chip Human Genome Array Strips were used for hybridisations. The genes that had altered expression levels in drug-resistant sublines were selected and filtered by scatter plots. The genes that were up- or downregulated more than fivefold were selected and listed. Among the investigated genes, 28 genes were upregulated, 10 genes were downregulated, and two genes were down- or upregulated depending on the cell line. Between upregulated genes 12 were upregulated very significantly—over 20-fold. These genes included COL1A2, COL12A1, COL21A1, LOX, TGFBI, LAMB1, EFEMP1, GPC3, SDC2, MGP, MMP3, and TIMP3. Four genes were very significantly downregulated: COL11A1, LAMA2, GPC6, and LUM. The expression profiles of investigated genes provide a preliminary insight into the relationship between drug resistance and the expression of ECM components. Identifying correlations between investigated genes and drug resistance will require further analysis. PMID:24804215

  8. Biomineralization of a Self-Assembled Extracellular Matrix for Bone Tissue Engineering

    SciTech Connect

    Yizhi, M.; Yi-Xian, Q; DiMasi, E; Xiaolan, B; Rafailovich, M; Pernodet, N

    2009-01-01

    Understanding how biomineralization occurs in the extracellular matrix (ECM) of bone cells is crucial to the understanding of bone formation and the development of a successfully engineered bone tissue scaffold. It is still unclear how ECM mechanical properties affect protein-mineral interactions in early stages of bone mineralization. We investigated the longitudinal mineralization properties of MC3T3-E1 cells and the elastic modulus of their ECM using shear modulation force microscopy, synchrotron grazing incidence X-ray diffraction (GIXD), scanning electron microscopy, energy dispersive X-ray spectroscopy, and confocal laser scanning microscopy (CLSM). The elastic modulus of the ECM fibers underwent significant changes for the mineralizing cells, which were not observed in the nonmineralizing cells. On substrates conducive to ECM network production, the elastic modulus of mineralizing cells increased at time points corresponding to mineral production, whereas that of the nonmineralizing cells did not vary over time. The presence of hydroxyapatite in mineralizing cells and the absence thereof in the nonmineralizing ones were confirmed by GIXD, and CLSM showed that a restructuring of actin occurred only for mineral-producing cells. These results show that the correct and complete development of the ECM network is required for osteoblasts to mineralize. This in turn requires a suitably prepared synthetic substrate for bone development to succeed in vitro.

  9. Cadherin-11 is a novel regulator of extracellular matrix synthesis and tissue mechanics.

    PubMed

    Row, Sindhu; Liu, Yayu; Alimperti, Stella; Agarwal, Sandeep K; Andreadis, Stelios T

    2016-08-01

    We discovered that Cadherin-11 (CDH11) regulates collagen and elastin synthesis, both affecting the mechanical properties and contractile function of animal tissues. Using a Cdh11-null mouse model, we observed a significant reduction in the mechanical properties [Youngs' modulus and ultimate tensile strength (UTS)] of Cdh11(-/-) as compared to wild-type (WT) mouse tissues, such as the aorta, bladder and skin. The deterioration of mechanical properties (Youngs' modulus and UTS) was accompanied by reduced collagen and elastin content in Cdh11(-/-) mouse tissues as well as in cells in culture. Similarly, knocking down CDH11 abolished collagen and elastin synthesis in human cells, and consequently reduced their ability to generate force. Conversely, engagement of CDH11 through homophilic interactions, led to swift activation of the TGF-β and ROCK pathways as evidenced by phosphorylation of downstream effectors. Subsequently, activation of the key transcription factors, MRTF-A (also known as MKL1) and MYOCD led to significant upregulation of collagen and elastin genes. Taken together, our results demonstrate a novel role of adherens junctions in regulating extracellular matrix (ECM) synthesis with implications for many important biological processes, including maintenance of tissue integrity, wound healing and tissue regeneration. PMID:27311482

  10. The Extracellular Matrix Protein Brevican Limits Time-Dependent Enhancement of Cocaine Conditioned Place Preference.

    PubMed

    Lubbers, Bart R; Matos, Mariana R; Horn, Annemarie; Visser, Esther; Van der Loo, Rolinka C; Gouwenberg, Yvonne; Meerhoff, Gideon F; Frischknecht, Renato; Seidenbecher, Constanze I; Smit, August B; Spijker, Sabine; van den Oever, Michel C

    2016-06-01

    Cocaine-associated environmental cues sustain relapse vulnerability by reactivating long-lasting memories of cocaine reward. During periods of abstinence, responding to cocaine cues can time-dependently intensify a phenomenon referred to as 'incubation of cocaine craving'. Here, we investigated the role of the extracellular matrix protein brevican in recent (1 day after training) and remote (3 weeks after training) expression of cocaine conditioned place preference (CPP). Wild-type and Brevican heterozygous knock-out mice, which express brevican at ~50% of wild-type levels, received three cocaine-context pairings using a relatively low dose of cocaine (5 mg/kg). In a drug-free CPP test, heterozygous mice showed enhanced preference for the cocaine-associated context at the remote time point compared with the recent time point. This progressive increase was not observed in wild-type mice and it did not generalize to contextual-fear memory. Virally mediated overexpression of brevican levels in the hippocampus, but not medial prefrontal cortex, of heterozygous mice prevented the progressive increase in cocaine CPP, but only when overexpression was induced before conditioning. Post-conditioning overexpression of brevican did not affect remote cocaine CPP, suggesting that brevican limited the increase in remote CPP by altering neuro-adaptive mechanisms during cocaine conditioning. We provide causal evidence that hippocampal brevican levels control time-dependent enhancement of cocaine CPP during abstinence, pointing to a novel substrate that regulates incubation of responding to cocaine-associated cues. PMID:26711251

  11. The role of extracellular matrix in age-related conduction disorders: a forgotten player?

    PubMed Central

    Spadaccio, Cristiano; Rainer, Alberto; Mozetic, Pamela; Trombetta, Marcella; Dion, Robert A; Barbato, Raffaele; Nappi, Francesco; Chello, Massimo

    2015-01-01

    Cardiovascular aging is a physiological process gradually leading to structural degeneration and functional loss of all the cardiac and vascular components. Conduction system is also deeply influenced by the aging process with relevant reflexes in the clinical side. Age-related arrhythmias carry significant morbidity and mortality and represent a clinical and economical burden. An important and unjustly unrecognized actor in the pathophysiology of aging is represented by the extracellular matrix (ECM) that not only structurally supports the heart determining its mechanical and functional properties, but also sends a biological signaling regulating cellular function and maintaining tissue homeostasis. At the biophysical level, cardiac ECM exhibits a peculiar degree of anisotropy, which is among the main determinants of the conductive properties of the specialized electrical conduction system. Age-associated alterations of cardiac ECM are therefore able to profoundly affect the function of the conduction system with striking impact on the patient clinical conditions. This review will focus on the ECM changes that occur during aging in the heart conduction system and on their translation to the clinical scenario. Potential diagnostic and therapeutical perspectives arising from the knowledge on ECM age-associated alterations are further discussed. PMID:25678907

  12. Intracellular Flow Cytometric Measurement of Extracellular Matrix Components in Porcine Intervertebral Disc Cells

    PubMed Central

    Flagler, Daniel J.; Huang, Chun-Yuh; Yuan, Tai-Yi; Lu, Zhongmin; Cheung, Herman S.; Gu, Wei Yong

    2009-01-01

    The objective of this study was to develop and demonstrate the utility of a novel method of evaluating intracellular levels of extracellular matrix (ECM) components in intervertebral disc (IVD) cells using flow cytometry. By using this method, this study discriminated between cell populations in porcine IVD and examined the response of IVD cells to monolayer cultures, a traditional method of cell expansion, by measuring phenotypic attributes of ECM component production. It was found that monolayer cultures affected collagen production of IVD cells while there were differences in collagen type II production between the cells isolated from the annulus fibrosus (AF) and nucleus pulposus (NP) regions of IVD. Size distributions of fresh and cultured cells were also presented while the relationships between cell size and intracellular collagen level revealed heterogeneous cell populations in AF and NP regions. Furthermore, this study showed that the intracellular collagen signals of IVD cells were significantly enhanced by the treatments of Brefeldin-A and ascorbic acid. This suggests that Brefeldin-A and ascorbic acid could be used to increase the sensitivity of flow cytometric analysis on intracellular collagen levels by maximizing collagen accumulation inside cells. Since a unique feature of the flow cytometric screening tool is the ability to discriminate between various cell populations in a single sample, the flow cytometric method developed in this study may have the potential to identify specific collagen-producing cell populations from tissues or cell cultures. PMID:20161070

  13. Activin A Suppresses Osteoblast Mineralization Capacity by Altering Extracellular Matrix (ECM) Composition and Impairing Matrix Vesicle (MV) Production*

    PubMed Central

    Alves, Rodrigo D. A. M.; Eijken, Marco; Bezstarosti, Karel; Demmers, Jeroen A. A.; van Leeuwen, Johannes P. T. M.

    2013-01-01

    During bone formation, osteoblasts deposit an extracellular matrix (ECM) that is mineralized via a process involving production and secretion of highly specialized matrix vesicles (MVs). Activin A, a transforming growth factor-β (TGF-β) superfamily member, was previously shown to have inhibitory effects in human bone formation models through unclear mechanisms. We investigated these mechanisms elicited by activin A during in vitro osteogenic differentiation of human mesenchymal stem cells (hMSC). Activin A inhibition of ECM mineralization coincided with a strong decline in alkaline phosphatase (ALP1) activity in extracellular compartments, ECM and matrix vesicles. SILAC-based quantitative proteomics disclosed intricate protein composition alterations in the activin A ECM, including changed expression of collagen XII, osteonectin and several cytoskeleton-binding proteins. Moreover, in activin A osteoblasts matrix vesicle production was deficient containing very low expression of annexin proteins. ECM enhanced human mesenchymal stem cell osteogenic development and mineralization. This osteogenic enhancement was significantly decreased when human mesenchymal stem cells were cultured on ECM produced under activin A treatment. These findings demonstrate that activin A targets the ECM maturation phase of osteoblast differentiation resulting ultimately in the inhibition of mineralization. ECM proteins modulated by activin A are not only determinant for bone mineralization but also possess osteoinductive properties that are relevant for bone tissue regeneration. PMID:23781072

  14. Effect of spaceflight on the extracellular matrix of skeletal muscle after a crush injury

    NASA Technical Reports Server (NTRS)

    Stauber, W. T.; Fritz, V. K.; Burkovskaia, T. E.; Il'ina-Kakueva, E. I.

    1992-01-01

    The organization and composition of the extracellular matrix were studied in the crush-injured gastrocnemius muscle of rats subjected to 0 G. After 14 days of flight on Cosmos 2044, the gastrocnemius muscle was removed and evaluated by histochemical and immunohistochemical techniques from the five injured flight rodents and various earth-based treatment groups. In general, the repair process was similar in all injured muscle samples with regard to the organization of the extracellular matrix and myofibers. Small and large myofibers were present within an expanded extracellular matrix, indicative of myogenesis and muscle regeneration. In the tail-suspended animals, a more complete repair was observed with nonenlarged area of nonmuscle cells or matrix material visible. In contrast, the muscle samples from the flight animals were less well organized and contained more macrophages and blood vessels in the repair region, indicative of a delayed repair process, but did not demonstrate any chronic inflammation. Myofiber repair did vary in muscles from the different groups, being slowest in the flight animals and most complete in the tail-suspended ones.

  15. A role for the extracellular matrix component hyaluronan in kidney dysfunction during ACE-inhibitor fetopathy.

    PubMed

    Hansell, P; Palm, F

    2015-04-01

    Despite data showing that inhibitors of the renin-angiotensin system increase the risks of fetal morbidity and dysfunctionality later in life, their use during pregnancy has increased. The fetopathy induced by angiotensin converting enzyme (ACE) inhibitors is characterized by anuria, hypotension and growth restriction, but can also be associated with pulmonary hypoplasia. In the kidney, this fetopathy includes atrophy of the medulla, reduced number of glomeruli, developmental lesions of tubules and vessels, tubulointerstitial inflammation and extracellular matrix accumulation. Although angiotensin II (Ang II) inhibition during nephrogenesis interferes with normal growth and development, this review will focus on effects of the heavily accumulated matrix component hyaluronan (HA). An important mechanism of HA accumulation during nephrogenesis is disruption of its normal reduction as a consequence of lack of Ang II activation of hyaluronidase. Hyaluronan has very large water-attracting properties and is pro-inflammatory when fragmented. The ensuing inflammation and interstitial oedema affect kidney function. Hyaluronan is colocalized with CD44 overexpression and infiltrating immune cells. These properties make HA a plausible contributor to the observed structural and functional kidney defects associated with the fetopathy. Available data support an involvement of HA in kidney dysfunction of the foetus and during adulthood due to the physico-chemical characteristics of HA. No clinical treatment for HA accumulation exists. Treatment with the HA-degrading enzyme hyaluronidase and an HA synthesis inhibitor has been tested successfully in experimental models in the kidney, heart and pancreas. Reduced HA accumulation to reduce interstitial oedema and inflammation may improve organ function, but this concept needs to be tested in a controlled study before causal relationships can be established. PMID:25600777

  16. The inhibitory effects of endostatin on endothelial cells are modulated by extracellular matrix

    SciTech Connect

    Delaney, Christie E.; Weagant, Brodie T.; Addison, Christina L. . E-mail: caddison@ohri.ca

    2006-08-01

    We investigated the ability of extracellular matrix (ECM) proteins to modulate the response of endothelial cells to both promoters and inhibitors of angiogenesis. Using human dermal microvascular endothelial cells (HDMEC), we found that cells demonstrated different adhesive properties and proliferative responses to the growth factor VEGF depending upon which ECM protein with which they were in contact, with fibronectin having the most impact on VEGF-induced HDMEC proliferation and survival. More importantly, we observed that ECM could modulate the ability of the angiogenic inhibitor endostatin to prevent endothelial cell proliferation, survival and migration. We observed that growth on vitronectin or fibronectin impaired the ability of endostatin to inhibit VEGF-induced HDMEC proliferation to the greatest extent as determined by BrdU incorporation. We found that, following growth on collagen I or collagen IV, endostatin only inhibited VEGF-induced HDMEC proliferation at the highest dose tested (2500 ng/ml). In a similar manner, we observed that growth on ECM proteins modulated the ability of endostatin to induce endothelial cell apoptosis, with growth on collagen I, fibronectin and collagen IV impairing endostatin-induced apoptosis. Interestingly, endostatin inhibited VEGF-induced HDMEC migration following culture on collagen I, collagen IV and laminin, while migration was not inhibited by endostatin following HDMEC culture on other matrices including vitronectin, fibronectin and tenascin-C. These results suggest that different matrix proteins may affect different mechanisms of endostatin inhibition of angiogenesis. Taken together, our results suggest that the ECM may have a profound impact on the ability of angiostatic molecules such as endostatin to inhibit angiogenesis and thus may have impact on the clinical efficacy of such inhibitors.

  17. Silicon-based quantum dots induce inflammation in human lung cells and disrupt extracellular matrix homeostasis.

    PubMed

    Stan, Miruna-Silvia; Sima, Cornelia; Cinteza, Ludmila Otilia; Dinischiotu, Anca

    2015-08-01

    Quantum dots (QDs) are nanocrystalline semiconductor materials that have been tested for biological applications such as cancer therapy, cellular imaging and drug delivery, despite the serious lack of information of their effects on mammalian cells. The present study aimed to evaluate the potential of Si/SiO2 QDs to induce an inflammatory response in MRC-5 human lung fibroblasts. Cells were exposed to different concentrations of Si/SiO2 QDs (25-200 μg·mL(-1)) for 24, 48, 72 and 96 h. The results obtained showed that uptake of QDs was dependent on biocorona formation and the stability of nanoparticles in various biological media (minimum essential medium without or with 10% fetal bovine serum). The cell membrane damage indicated by the increase in lactate dehydrogenase release after exposure to QDs was dose- and time-dependent. The level of lysosomes increased proportionally with the concentration of QDs, whereas an accumulation of autophagosomes was also observed. Cellular morphology was affected, as shown by the disruption of actin filaments. The enhanced release of nitric oxide and the increase in interleukin-6 and interleukin-8 protein expression suggested that nanoparticles triggered an inflammatory response in MRC-5 cells. QDs decreased the protein expression and enzymatic activity of matrix metalloproteinase (MMP)-2 and MMP-9 and also MMP-1 caseinase activity, whereas the protein levels of MMP-1 and tissue inhibitor of metalloproteinase-1 increased. The present study reveals for the first time that silicon-based QDs are able to generate inflammation in lung cells and cause an imbalance in extracellular matrix turnover through a differential regulation of MMPs and tissue inhibitor of metalloproteinase-1 protein expression. PMID:26032556

  18. Redox-Relevant Aspects of the Extracellular Matrix and Its Cellular Contacts via Integrins

    PubMed Central

    de Rezende, Flávia Figueiredo

    2014-01-01

    Abstract Significance: The extracellular matrix (ECM) fulfills essential functions in multicellular organisms. It provides the mechanical scaffold and environmental cues to cells. Upon cell attachment, the ECM signals into the cells. In this process, reactive oxygen species (ROS) are physiologically used as signalizing molecules. Recent Advances: ECM attachment influences the ROS-production of cells. In turn, ROS affect the production, assembly and turnover of the ECM during wound healing and matrix remodeling. Pathological changes of ROS levels lead to excess ECM production and increased tissue contraction in fibrotic disorders and desmoplastic tumors. Integrins are cell adhesion molecules which mediate cell adhesion and force transmission between cells and the ECM. They have been identified as a target of redox-regulation by ROS. Cysteine-based redox-modifications, together with structural data, highlighted particular regions within integrin heterodimers that may be subject to redox-dependent conformational changes along with an alteration of integrin binding activity. Critical Issues: In a molecular model, a long-range disulfide-bridge within the integrin β-subunit and disulfide bridges within the genu and calf-2 domains of the integrin α-subunit may control the transition between the bent/inactive and upright/active conformation of the integrin ectodomain. These thiol-based intramolecular cross-linkages occur in the stalk domain of both integrin subunits, whereas the ligand-binding integrin headpiece is apparently unaffected by redox-regulation. Future Directions: Redox-regulation of the integrin activation state may explain the effect of ROS in physiological processes. A deeper understanding of the underlying mechanism may open new prospects for the treatment of fibrotic disorders. Antioxid. Redox Signal. 20, 1977–1993. PMID:24040997

  19. Leptospira interrogans induces uterine inflammatory responses and abnormal expression of extracellular matrix proteins in dogs.

    PubMed

    Wang, Wei; Gao, Xuejiao; Guo, Mengyao; Zhang, Wenlong; Song, Xiaojing; Wang, Tiancheng; Zhang, Zecai; Jiang, Haichao; Cao, Yongguo; Zhang, Naisheng

    2014-10-01

    Leptospira interrogans (L. interrogans), a worldwide zoonosis, infect humans and animals. In dogs, four syndromes caused by leptospirosis have been identified: icteric, hemorrhagic, uremic (Stuttgart disease) and reproductive (abortion and premature or weak pups), and also it caused inflammation. Extracellular matrix (ECM) is a complex mixture of matrix molecules that is crucial to the reproduction. Both inflammatory response and ECM are closed relative to reproductive. The aim of this study was to clarify how L. interrogans affected the uterus of dogs, by focusing on the inflammatory responses, and ECM expression in dogs uterine tissue infected by L. interrogans. In the present study, 27 dogs were divided into 3 groups, intrauterine infusion with L. interrogans, to make uterine infection, sterile EMJH, and normal saline as a control, respectively. The uteruses were removed by surgical operation in 10, 20, and 30 days, respectively. The methods of histopathological analysis, ELISA, Western blot and qPCR were used. The results showed that L. interrogans induced significantly inflammatory responses, which were characterized by inflammatory cellular infiltration and high expression levels of tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) in uterine tissue of these dogs. Furthermore, L. interrogans strongly down-regulated the expression of ECM (collagens (CL) IV, fibronectins (FN) and laminins (LN)) in mRNA and protein levels. These data indicated that strongly inflammatory responses, and abnormal regulation of ECM might contribute to the proliferation of dogs infected by L. interrogans. PMID:25153777

  20. Sum of the Parts: Composition and Architecture of the Bacterial Extracellular Matrix

    PubMed Central

    McCrate, Oscar A.; Zhou, Xiaoxue; Reichhardt, Courtney; Cegelski, Lynette

    2013-01-01

    Bacterial biofilms are complex multicellular assemblies that exhibit resistance to antibiotics and contribute to the pathogenesis of serious and chronic infectious diseases. New approaches and quantitative data are needed to define the molecular composition of bacterial biofilms. E. coli biofilms are known to contain polysaccharides and functional amyloid fibers termed curli, yet accurate determinations of biofilm composition at the molecular level have been elusive. The ability to define the composition of the extracellular matrix is crucial for the elucidation of structure–function relationships that will aid the development of chemical strategies to disrupt biofilms. We have developed an approach that integrates non-perturbative preparation of the extracellular matrix with electron microscopy, biochemistry and solid-state NMR spectroscopy to define the chemical composition of the intact and insoluble extracellular matrix of a clinically important pathogenic bacterium—uropathogenic E. coli. Our data permitted a sum-of-all-the-parts analysis. Electron microscopy revealed supramolecular shell-like structures that encapsulated single cells and enmeshed the bacterial community. Biochemical and solid-state NMR measurements of the matrix and constitutive parts established that the matrix is composed of two major components, curli and cellulose, each in a quantifiable amount. This approach to quantifying the matrix composition is widely applicable to other organisms and to examining the influence of biofilm inhibitors. Collectively, our NMR spectra and the electron micrographs of the purified ECM inspire us to consider the biofilm matrix not as an undefined slime, but as an assembly of polymers with a defined composition and architecture. PMID:23827139

  1. Extracellular Matrix Stiffness Controls VEGF Signaling and Processing in Endothelial Cells.

    PubMed

    Sack, Kelsey D; Teran, Madelane; Nugent, Matthew A

    2016-09-01

    Vascular endothelial growth factor A (VEGF) drives endothelial cell maintenance and angiogenesis. Endothelial cell behavior is altered by the stiffness of the substrate the cells are attached to suggesting that VEGF activity might be influenced by the mechanical cellular environment. We hypothesized that extracellular matrix (ECM) stiffness modifies VEGF-cell-matrix tethering leading to altered VEGF processing and signaling. We analyzed VEGF binding, internalization, and signaling as a function of substrate stiffness in endothelial cells cultured on fibronectin (Fn) linked polyacrylamide gels. Cell produced extracellular matrices on the softest substrates were least capable of binding VEGF, but the cells exhibited enhanced VEGF internalization and signaling compared to cells on all other substrates. Inhibiting VEGF-matrix binding with sucrose octasulfate decreased cell-internalization of VEGF and, inversely, heparin pre-treatment to enhance Fn-matrix binding of VEGF increased cell-internalization of VEGF regardless of matrix stiffness. β1 integrins, which connect cells to Fn, modulated VEGF uptake in a stiffness dependent fashion. Cells on hard surfaces showed decreased levels of activated β1 and inhibition of β1 integrin resulted in a greater proportional decrease in VEGF internalization than in cells on softer matrices. Extracellular matrix binding is necessary for VEGF internalization. Stiffness modifies the coordinated actions of VEGF-matrix binding and β1 integrin binding/activation, which together are critical for VEGF internalization. This study provides insight into how the microenvironment may influence tissue regeneration and response to injury and disease. J. Cell. Physiol. 231: 2026-2039, 2016. © 2016 Wiley Periodicals, Inc. PMID:26773314

  2. Heterogeneous expression of extracellular matrix molecules in the red nucleus of the rat.

    PubMed

    Rácz, É; Gaál, B; Matesz, C

    2016-05-13

    Previous studies in our laboratory showed that the organization and heterogeneous molecular composition of extracellular matrix is associated with the variable cytoarchitecture, connections and specific functions of the vestibular nuclei and two related areas of the vestibular neural circuits, the inferior olive and prepositus hypoglossi nucleus. The aim of the present study is to reveal the organization and distribution of various molecular components of extracellular matrix in the red nucleus, a midbrain premotor center. Morphologically and functionally the red nucleus is comprised of the magno- and parvocellular parts, with overlapping neuronal population. By using histochemical and immunohistochemical methods, the extracellular matrix appeared as perineuronal net, axonal coat, perisynaptic matrix or diffuse network in the neuropil. In both parts of the red nucleus we have observed positive hyaluronan, tenascin-R, link protein, and lectican (aggrecan, brevican, versican, neurocan) reactions. Perineuronal nets were detected with each of the reactions and the aggrecan showed the most intense staining in the pericellular area. The two parts were clearly distinguished on the basis of neurocan and HAPLN1 expression as they have lower intensity in the perineuronal nets of large cells and in the neuropil of the magnocellular part. Additionally, in contrast to this pattern, the aggrecan was heavily labeled in the magnocellular region sharply delineating from the faintly stained parvocellular area. The most characteristic finding was that the appearance of perineuronal nets was related with the neuronal size independently from its position within the two subdivisions of red nucleus. In line with these statements none of the extracellular matrix molecules were restricted exclusively to the magno- or parvocellular division. The chemical heterogeneity of the perineuronal nets may support the recently accepted view that the red nucleus comprises more different populations of

  3. Extracellular matrix-associated proteins form an integral and dynamic system during Pseudomonas aeruginosa biofilm development

    PubMed Central

    Zhang, Weipeng; Sun, Jin; Ding, Wei; Lin, Jinshui; Tian, Renmao; Lu, Liang; Liu, Xiaofen; Shen, Xihui; Qian, Pei-Yuan

    2015-01-01

    Though the essential role of extracellular matrix in biofilm development has been extensively documented, the function of matrix-associated proteins is elusive. Determining the dynamics of matrix-associated proteins would be a useful way to reveal their functions in biofilm development. Therefore, we applied iTRAQ-based quantitative proteomics to evaluate matrix-associated proteins isolated from different phases of Pseudomonas aeruginosa ATCC27853 biofilms. Among the identified 389 proteins, 54 changed their abundance significantly. The increased abundance of stress resistance and nutrient metabolism-related proteins over the period of biofilm development was consistent with the hypothesis that biofilm matrix forms micro-environments in which cells are optimally organized to resist stress and use available nutrients. Secreted proteins, including novel putative effectors of the type III secretion system were identified, suggesting that the dynamics of pathogenesis-related proteins in the matrix are associated with biofilm development. Interestingly, there was a good correlation between the abundance changes of matrix-associated proteins and their expression. Further analysis revealed complex interactions among these modulated proteins, and the mutation of selected proteins attenuated biofilm development. Collectively, this work presents the first dynamic picture of matrix-associated proteins during biofilm development, and provides evidences that the matrix-associated proteins may form an integral and well regulated system that contributes to stress resistance, nutrient acquisition, pathogenesis and the stability of the biofilm. PMID:26029669

  4. Soil organic matter and the extracellular microbial matrix show contrasting responses to C and N availability

    PubMed Central

    Redmile-Gordon, M.A.; Evershed, R.P.; Hirsch, P.R.; White, R.P.; Goulding, K.W.T.

    2015-01-01

    An emerging paradigm in soil science suggests microbes can perform ‘N mining’ from recalcitrant soil organic matter (SOM) in conditions of low N availability. However, this requires the production of extracellular structures rich in N (including enzymes and structural components) and thus defies stoichiometric expectation. We set out to extract newly synthesised peptides from the extracellular matrix in soil and compare the amino acid (AA) profiles, N incorporation and AA dynamics in response to labile inputs of contrasting C/N ratio. Glycerol was added both with and without an inorganic source of N (10% 15N labelled NH4NO3) to a soil already containing a large pool of refractory SOM and incubated for 10 days. The resulting total soil peptide (TSP) and extracellular pools were compared using colorimetric methods, gas chromatography, and isotope ratio mass spectrometry. N isotope compositions showed that the extracellular polymeric substance (EPS) contained a greater proportion of products formed de novo than did TSP, with hydrophobic EPS-AAs (leucine, isoleucine, phenylalanine, hydroxyproline and tyrosine) deriving substantially more N from the inorganic source provided. Quantitative comparison between extracts showed that the EPS contained greater relative proportions of alanine, glycine, proline, phenylalanine and tyrosine. The greatest increases in EPS-peptide and EPS-polysaccharide concentrations occurred at the highest C/N ratios. All EPS-AAs responded similarly to treatment whereas the responses of TSP were more complex. The results suggest that extracellular investment of N (as EPS peptides) is a microbial survival mechanism in conditions of low N/high C which, from an evolutionary perspective, must ultimately lead to the tendency for increased N returns to the microbial biomass. A conceptual model is proposed that describes the dynamics of the extracellular matrix in response to the C/N ratio of labile inputs. PMID:26339106

  5. Endocytosis of collagen by hepatic stellate cells regulates extracellular matrix dynamics

    PubMed Central

    Bi, Yan; Mukhopadhyay, Dhriti; Drinane, Mary; Ji, Baoan; Li, Xing; Cao, Sheng

    2014-01-01

    Hepatic stellate cells (HSCs) generate matrix, which in turn may also regulate HSCs function during liver fibrosis. We hypothesized that HSCs may endocytose matrix proteins to sense and respond to changes in microenvironment. Primary human HSCs, LX2, or mouse embryonic fibroblasts (MEFs) [wild-type; c-abl−/−; or Yes, Src, and Fyn knockout mice (YSF−/−)] were incubated with fluorescent-labeled collagen or gelatin. Fluorescence-activated cell sorting analysis and confocal microscopy were used for measuring cellular internalization of matrix proteins. Targeted PCR array and quantitative real-time PCR were used to evaluate gene expression changes. HSCs and LX2 cells endocytose collagens in a concentration- and time-dependent manner. Endocytosed collagen colocalized with Dextran 10K, a marker of macropinocytosis, and 5-ethylisopropyl amiloride, an inhibitor of macropinocytosis, reduced collagen internalization by 46%. Cytochalasin D and ML7 blocked collagen internalization by 47% and 45%, respectively, indicating that actin and myosin are critical for collagen endocytosis. Wortmannin and AKT inhibitor blocked collagen internalization by 70% and 89%, respectively, indicating that matrix macropinocytosis requires phosphoinositide-3-kinase (PI3K)/AKT signaling. Overexpression of dominant-negative dynamin-2 K44A blocked matrix internalization by 77%, indicating a role for dynamin-2 in matrix macropinocytosis. Whereas c-abl−/− MEF showed impaired matrix endocytosis, YSF−/− MEF surprisingly showed increased matrix endocytosis. It was also associated with complex gene regulations that related with matrix dynamics, including increased matrix metalloproteinase 9 (MMP-9) mRNA levels and zymographic activity. HSCs endocytose matrix proteins through macropinocytosis that requires a signaling network composed of PI3K/AKT, dynamin-2, and c-abl. Interaction with extracellular matrix regulates matrix dynamics through modulating multiple gene expressions including MMP-9

  6. Immunohistochemical study of cytoskeletal and extracellular matrix components in the notochord and notochordal sheath of amphioxus

    PubMed Central

    Bočina, Ivana; Saraga-Babić, Mirna

    2006-01-01

    A major cytoskeletal and extracellular matrix proteins of the amphioxus notochordal cells and sheath were detected by immunohistochemical techniques. The three-layered amphioxus notochordal sheath strongly expressed fish collagen type I in its outer and middle layers, while in the innermost layer expression did not occur. The amphioxus notochordal sheath was reactive to applied anti-human antibodies for intermediate filament proteins such as cytokeratins, desmin and vimentin, as well as to microtubule components (ß-tubulin), particularly in the area close to the epipharyngeal groove. Alpha-smooth muscle actin was expressed in some notochordal cells and in the area of the notochordal attachment to the sheath. Thus muscular nature of notochordal cells was shown by immunohistochemistry in tissue section. Our results confirm that genes encoding intermediate filament proteins, microtubules and microfilaments are highly conserved during evolution. Collagen type I was proven to be the key extracellular matrix protein that forms the amphioxus notochordal sheath. PMID:16733537

  7. How Osteoblasts Sense their Environment: Integrin-Extracellular Matrix Interactions and Mechanical Loading of Bone

    NASA Technical Reports Server (NTRS)

    Globus, Ruth K.; Dalton, Bonnie (Technical Monitor)

    2002-01-01

    Osteoblasts are the cells responsible for forming and replacing bone throughout life. We know that mechanical stimulation through weight-bearing at I gravity on Earth is needed to maintain healthy bone, and that osteoblasts play a critical role in that process. Over the last 9 years in my laboratory at NASA ARC, we have studied the regulation of osteoblast function by interactions between the extracellular matrix and die cell. Using a cell culture approach, we defined the repertoire of adhesion receptors, called integrins, which are expressed on the osteoblast surface, as well as specific extracellular matrix proteins, which are needed for cellular differentiation and survival. We are now extending these observations to determine if integrin signaling is involved in the skeletal responses to disuse and recovery from disuse using the rodent model of hindlimb unloading by tail suspension. Together, our cell culture and animal studies are providing new insight into the regulation of osteoblast function in bone.

  8. Designer Extracellular Matrix Based on DNA-Peptide Networks Generated by Polymerase Chain Reaction.

    PubMed

    Finke, Alexander; Bußkamp, Holger; Manea, Marilena; Marx, Andreas

    2016-08-16

    Cell proliferation and differentiation in multicellular organisms are partially regulated by signaling from the extracellular matrix. The ability to mimic an extracellular matrix would allow particular cell types to be specifically recognized, which is central to tissue engineering. We present a new functional DNA-based material with cell-adhesion properties. It is generated by using covalently branched DNA as primers in PCR. These primers were functionalized by click chemistry with the cyclic peptide c(RGDfK), a peptide that is known to predominantly bind to αvβ3 integrins, which are found on endothelial cells and fibroblasts, for example. As a covalent coating of surfaces, this DNA-based material shows cell-repellent properties in its unfunctionalized state and gains adhesiveness towards specific target cells when functionalized with c(RGDfK). These cells remain viable and can be released under mild conditions by DNase I treatment. PMID:27410200

  9. Chondroitinase injection improves keloid pathology by reorganizing the extracellular matrix with regenerated elastic fibers.

    PubMed

    Ishiko, Toshihiro; Naitoh, Motoko; Kubota, Hiroshi; Yamawaki, Satoko; Ikeda, Mika; Yoshikawa, Katsuhiro; Fujita, Hiroshi; Yamaguchi, Hiroaki; Kurahashi, Yasuhiro; Suzuki, Shigehiko

    2013-05-01

    Keloids are a proliferative fibrotic disease characterized by abnormal accumulation of extracellular matrix in the dermis. Keloid lesions lack skin plasticity due to deficiencies in elastic fiber formation in the extracellular matrix. The loss of elastic fiber is caused by excessive accumulation of chondroitin sulfate (CS), a sulfated glycosaminoglycan. However, there is no radical cure for keloids. Using a model system, we show herein that treatment of keloid tissues with chondroitinase ABC, an enzyme that specifically digests CS, improves clinical features of keloids. Keloid tissues obtained from patients were grafted on nude mice, and chondroitinase ABC was injected into the grafted keloid tissues. Chondroitinase ABC treatment significantly reduced the volume of keloid implants concomitant with recovery of elastic fiber formation. These results suggest that chondroitinase ABC injection is an effective therapy for keloid. PMID:23451938

  10. Regulation of Extracellular Matrix Remodeling Proteins by Osteoblasts in Titanium Nanoparticle-Induced Aseptic Loosening Model.

    PubMed

    Xie, Jing; Hou, Yanhua; Fu, Na; Cai, Xiaoxiao; Li, Guo; Peng, Qiang; Lin, Yunfeng

    2015-10-01

    Titanium (Ti)-wear particles, formed at the bone-implant interface, are responsible for aseptic loosening, which is a main cause of total joint replacement failure. There have been many studies on Ti particle-induced function changes in mono-cultured osteoblasts and synovial cells. However, little is known on extracellular matrix remodeling displayed by osteoblasts when in coexistence with Synovial cells. To further mimic the bone-implant interface environment, we firstly established a nanoscaled-Ti particle-induced aseptic loosening system by co-culturing osteoblasts and Synovial cells. We then explored the impact of the Synovial cells on Ti particle-engulfed osteoblasts in the mimicked flamed niche. The matrix metalloproteinases and lysyl oxidases expression levels, two protein families which are critical in osseointegration, were examined under induction by tumor necrosis factor-alpha. It was found that the co-culture between the osteoblasts and Synovial cells markedly increased the migration and proliferation of the osteoblasts, even in the Ti-particle engulfed osteoblasts. Importantly, the Ti-particle engulfed osteoblasts, induced by TNF-alpha after the co-culture, enhanced the release of the matrix metalloproteinases and reduced the expressions of lysyl oxidases. The regulation of extracellular matrix remodeling at the protein level was further assessed by investigations on gene expression of the matrix metalloproteinases and lysyl oxidases, which also suggested that the regulation started at the genetic level. Our research work has therefore revealed the critical role of multi cell-type interactions in the extracellular matrix remodeling within the peri-prosthetic tissues, which provides new insights on aseptic loosening and brings new clues about incomplete osseointegration between the implantation materials and their surrounding bones. PMID:26502645

  11. Immunohistochemical evidence of rapid extracellular matrix remodeling after iron-particle irradiation of mouse mammary gland

    NASA Technical Reports Server (NTRS)

    Ehrhart, E. J.; Gillette, E. L.; Barcellos-Hoff, M. H.; Chaterjee, A. (Principal Investigator)

    1996-01-01

    High-LET radiation has unique physical and biological properties compared to sparsely ionizing radiation. Recent studies demonstrate that sparsely ionizing radiation rapidly alters the pattern of extracellular matrix expression in several tissues, but little is known about the effect of heavy-ion radiation. This study investigates densely ionizing radiation-induced changes in extracellular matrix localization in the mammary glands of adult female BALB/c mice after whole-body irradiation with 0.8 Gy 600 MeV iron particles. The basement membrane and interstitial extracellular matrix proteins of the mammary gland stroma were mapped with respect to time postirradiation using immunofluorescence. Collagen III was induced in the adipose stroma within 1 day, continued to increase through day 9 and was resolved by day 14. Immunoreactive tenascin was induced in the epithelium by day 1, was evident at the epithelial-stromal interface by day 5-9 and persisted as a condensed layer beneath the basement membrane through day 14. These findings parallel similar changes induced by gamma irradiation but demonstrate different onset and chronicity. In contrast, the integrity of epithelial basement membrane, which was unaffected by sparsely ionizing radiation, was disrupted by iron-particle irradiation. Laminin immunoreactivity was mildly irregular at 1 h postirradiation and showed discontinuities and thickening from days 1 to 9. Continuity was restored by day 14. Thus high-LET radiation, like sparsely ionizing radiation, induces rapid-remodeling of the stromal extracellular matrix but also appears to alter the integrity of the epithelial basement membrane, which is an important regulator of epithelial cell proliferation and differentiation.

  12. Prion permissive pathways: extracellular matrix genes control susceptibility to prion infection

    PubMed Central

    Imberdis, Thibaut; Harris, David A

    2014-01-01

    There are wide variations in the susceptibility of humans, animals, and cultured cell lines to infection by prions. In this issue of The EMBO Journal, Marbiah et al (2014) identified a gene regulatory network that regulates the susceptibility of cultured cells to prion infection. Surprisingly, a number of these genes impact the structure of the extracellular matrix. These results have important implications for understanding mechanisms of prion infection and also suggest new therapeutic targets. PMID:24952893

  13. Immunohistochemical evidence of rapid extracellular matrix remodeling after iron-particle irradiation of mouse mammary gland

    SciTech Connect

    Ehrhart, E.J.; Gillette, E.L.; Barcellos-Hoff, M.H.

    1996-02-01

    High-LET radiation has unique physical and biological properties compared to sparsely ionizing radiation. Recent studies demonstrate that sparsely ionizing radiation rapidly alters the pattern of extracellular matrix expression in several tissues, but little is known about the effect of heavy-ion radiation. This study investigates densely ionizing radiation-induced changes in extracellular matrix localization in the mammary glands of adult female BALB/c mice after whole-body irradiation with 0.8 Gy 600 MeV iron particles. The basement membrane and interstitial extracellular matrix proteins of the mammary gland stroma were mapped with respect to time postirradiation using immunofluorescence. Collagen III was induced in the adipose stroma within 1 day, continued to increase through day 9 and was resolved by day 14. Immunoreactive tenascin was induced in the epithelium by day 1, was evident at the epithelial-stromal interface by day 5-9 and persisted as a condensed layer beneath the basement membrane through day 14. These findings parallel similar changes induced by {gamma} irradiation but demonstrate different onset and chronicity. In contrast, the integrity of epithelial basement membrane, which was unaffected by sparsely ionizing radiation, was disrupted by iron-particle irradiation. Laminin inummoreactivity was mildly irregular at 1 h postirradiation and showed discontinuities and thickening from days 1 to 9. Continuity was restored by day 14. Thus high-LET radiation, like sparsely ionizing radiation, induces rapid remodeling of the stromal extracellular matrix but also appears to alter the integrity of the epithelial basement membrane, which is an important regulator of epithelial cell proliferation and differentiation. 40 refs., 3 figs.

  14. An Assay to Quantify Chemotactic Properties of Degradation Products from Extracellular Matrix

    PubMed Central

    Sicari, Brian M.; Zhang, Li; Londono, Ricardo; Badylak, Stephen F.

    2015-01-01

    The endogenous chemotaxis of cells toward sites of tissue injury and/or biomaterial implantation is an important component of the host response. Implanted biomaterials capable of recruiting host stem/progenitor cells to a site of interest may obviate challenges associated with cell transplantation. An assay for the identification and quantification of chemotaxis induced by surgically placed biologic scaffolds composed of extracellular matrix is described herein. PMID:24155230

  15. Bral1: "Superglue" for the extracellular matrix in the brain white matter.

    PubMed

    Cicanic, Michal; Sykova, Eva; Vargova, Lydia

    2012-04-01

    Bral1 is a link protein that stabilizes the binding between lecticans and hyaluronic acid and thus maintains the extracellular matrix assembly in the CNS. Bral1 is specifically located in the white matter around the nodes of Ranvier. Recent studies suggest its function in promoting saltatory neural conduction. This article reviews the current knowledge about the structure, expression and function of this link protein. PMID:22300985

  16. The ECM-Cell Interaction of Cartilage Extracellular Matrix on Chondrocytes

    PubMed Central

    Liu, Shuyun; Huang, Jingxiang; Guo, Weimin; Chen, Jifeng; Zhang, Li; Zhao, Bin; Peng, Jiang; Wang, Aiyuan; Wang, Yu; Xu, Wenjing; Lu, Shibi; Yuan, Mei; Guo, Quanyi

    2014-01-01

    Cartilage extracellular matrix (ECM) is composed primarily of the network type II collagen (COLII) and an interlocking mesh of fibrous proteins and proteoglycans (PGs), hyaluronic acid (HA), and chondroitin sulfate (CS). Articular cartilage ECM plays a crucial role in regulating chondrocyte metabolism and functions, such as organized cytoskeleton through integrin-mediated signaling via cell-matrix interaction. Cell signaling through integrins regulates several chondrocyte functions, including differentiation, metabolism, matrix remodeling, responses to mechanical stimulation, and cell survival. The major signaling pathways that regulate chondrogenesis have been identified as wnt signal, nitric oxide (NO) signal, protein kinase C (PKC), and retinoic acid (RA) signal. Integrins are a large family of molecules that are central regulators in multicellular biology. They orchestrate cell-cell and cell-matrix adhesive interactions from embryonic development to mature tissue function. In this review, we emphasize the signaling molecule effect and the biomechanics effect of cartilage ECM on chondrogenesis. PMID:24959581

  17. Age-related disruption of autophagy in dermal fibroblasts modulates extracellular matrix components

    SciTech Connect

    Tashiro, Kanae; Shishido, Mayumi; Fujimoto, Keiko; Hirota, Yuko; Yo, Kazuyuki; Gomi, Takamasa; Tanaka, Yoshitaka

    2014-01-03

    Highlights: •Autophagosomes accumulate in aged dermal fibroblasts. •Autophagic degradation is impaired in aged dermal fibroblasts. •Autophagy disruption affects extracellular matrix components in dermal fibroblasts. -- Abstract: Autophagy is an intracellular degradative system that is believed to be involved in the aging process. The contribution of autophagy to age-related changes in the human skin is unclear. In this study, we examined the relationship between autophagy and skin aging. Transmission electron microscopy and immunofluorescence microscopy analyses of skin tissue and cultured dermal fibroblasts derived from women of different ages revealed an increase in the number of nascent double-membrane autophagosomes with age. Western blot analysis showed that the amount of LC3-II, a form associated with autophagic vacuolar membranes, was significantly increased in aged dermal fibroblasts compared with that in young dermal fibroblasts. Aged dermal fibroblasts were minimally affected by inhibition of autophagic activity. Although lipofuscin autofluorescence was elevated in aged dermal fibroblasts, the expression of Beclin-1 and Atg5—genes essential for autophagosome formation—was similar between young and aged dermal fibroblasts, suggesting that the increase of autophagosomes in aged dermal fibroblasts was due to impaired autophagic flux rather than an increase in autophagosome formation. Treatment of young dermal fibroblasts with lysosomal protease inhibitors, which mimic the condition of aged dermal fibroblasts with reduced autophagic activity, altered the fibroblast content of type I procollagen, hyaluronan and elastin, and caused a breakdown of collagen fibrils. Collectively, these findings suggest that the autophagy pathway is impaired in aged dermal fibroblasts, which leads to deterioration of dermal integrity and skin fragility.

  18. Extracellular matrix components induce endocrine differentiation in vitro in NCI-H716 cells.

    PubMed Central

    de Bruïne, A. P.; Dinjens, W. N.; van der Linden, E. P.; Pijls, M. M.; Moerkerk, P. T.; Bosman, F. T.

    1993-01-01

    Endocrine cells occur in +/- 30% of colorectal adenocarcinomas. The significance of this phenomenon in terms of tumor behavior is still controversial. Endocrine differentiation in colorectal cancer cell lines is almost confined to tumor xenografts in vivo, suggesting that endocrine differentiation might be regulated by epithelial-stromal interactions. This hypothesis was studied in the cecal adenocarcinoma-derived cell line NCI-H716 by comparing the expression of chromogranin A protein and messenger RNA in vivo and in vitro and by attempts to induce differentiation in vitro. We found that chromogranin A expression, which was strongest in vivo, could be significantly enhanced in vitro by culturing tumor cells in the presence of native extracellular matrix, on fibroblast feeder layers, and in a defined medium with basic fibroblast growth factor. The results suggest that the extracellular matrix induces endocrine differentiation through factors (e.g., basic fibroblast-growth factor) that may be produced by stromal cells and after secretion bind to the extracellular matrix. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 6 PMID:8456938

  19. N-terminal specific conjugation of extracellular matrix proteins to 2-pyridinecarboxaldehyde functionalized polyacrylamide hydrogels.

    PubMed

    Lee, Jessica P; Kassianidou, Elena; MacDonald, James I; Francis, Matthew B; Kumar, Sanjay

    2016-09-01

    Polyacrylamide hydrogels have been used extensively to study cell responses to the mechanical and biochemical properties of extracellular matrix substrates. A key step in fabricating these substrates is the conjugation of cell adhesion proteins to the polyacrylamide surfaces, which typically involves nonspecifically anchoring these proteins via side-chain functional groups. This can result in a loss of presentation control and altered bioactivity. Here, we describe a new functionalization strategy in which we anchor full-length extracellular matrix proteins to polyacrylamide substrates using 2-pyridinecarboxaldehyde, which can be co-polymerized into polyacrylamide gels and used to immobilize proteins by their N-termini. This one-step reaction proceeds under mild aqueous conditions and does not require additional reagents. We demonstrate that these substrates can readily conjugate to various extracellular matrix proteins, as well as promote cell adhesion and spreading. Notably, this chemistry supports the assembly and cellular remodeling of large collagen fibers, which is not observed using conventional side-chain amine-conjugation chemistry. PMID:27348850

  20. Quantitation and relative distribution of extracellular matrix in Staphylococcus epidermidis biofilm

    SciTech Connect

    Van Pett, K.; Schurman, D.J.; Smith, R.L. )

    1990-05-01

    The relationship between adherence of bacteria to foreign bodies and their deposition of extracellular matrix was examined on glass and suture material. To quantitate bacterial adherence, uptake of ({sup 3}H)thymidine into bacterial DNA was analyzed. Corresponding amounts of extracellular matrix were measured by a new technique using ({sup 14}C)glucose incorporation. This study shows that ({sup 14}C)glucose preferentially labeled bacterial strains in proportion to biofilm production. The ratio of {sup 3}H{sup 14}C in high biofilm producers was 0.9 and in low producers it was 3.7. Radioactive identification of organisms as high and low producers was confirmed by electron microscopy. The results presented here show that production and accumulation of biofilm over time is a stable characteristic in different strains of S. epidermidis. The use of ratios reflecting radiolabeling of bacteria and biofilm by ({sup 3}H)thymidine and ({sup 14}C)glucose, respectively, is a quantitative yet simple technique to assess extracellular matrix of different strains of S. epidermidis.

  1. Role of Microvascular Tone and Extracellular Matrix Contraction in the Regulation of Interstitial Fluid: Implications for Aortic Dissection.

    PubMed

    Mallat, Ziad; Tedgui, Alain; Henrion, Daniel

    2016-09-01

    The pathophysiology of aortic dissection is poorly understood, and its risk is resistant to medical treatment. Most studies have focused on a proposed pathogenic role of transforming growth factor-β in Marfan disease and related thoracic aortic aneurysms and aortic dissections. However, clinical testing of this concept using angiotensin II type 1 receptor antagonists to block transforming growth factor-β signaling fell short of promise. Genetic mutations that predispose to thoracic aortic aneurysms and aortic dissections affect components of the extracellular matrix and proteins involved in cellular force generation. Thus, a role for dysfunctional mechanosensing in abnormal aortic wall remodeling is emerging. However, how abnormal mechanosensing leads to aortic dissection remains a mystery. Here, we review current knowledge about the regulation of interstitial fluid dynamics and myogenic tone and propose that alteration in contractile force reduces vascular tone in the microcirculation (here, aortic vasa vasorum) and leads to elevations of blood flow, transmural pressure, and fluid flux into the surrounding aortic media. Furthermore, reduced contractile force in medial smooth muscle cells coupled with alteration of structural components of the extracellular matrix limits extracellular matrix contraction, further promoting the formation of intramural edema, a critical step in the initiation of aortic dissection. The concept is supported by several pathophysiological and clinical observations. A direct implication of this concept is that drugs that lower blood pressure and limit interstitial fluid accumulation while preserving or increasing microvascular tone would limit the risk of dissection. In contrast, drugs that substantially lower microvascular tone would be ineffective or may accelerate the disease and precipitate aortic dissection. PMID:27444198

  2. Dynamic interactions between cells and their extracellular matrix mediate embryonic development.

    PubMed

    Goody, Michelle F; Henry, Clarissa A

    2010-06-01

    Cells and their surrounding extracellular matrix microenvironment interact throughout all stages of life. Understanding the continuously changing scope of cell-matrix interactions in vivo is crucial to garner insights into both congenital birth defects and disease progression. A current challenge in the field of developmental biology is to adapt in vitro tools and rapidly evolving imaging technology to study cell-matrix interactions in a complex 4-D environment. In this review, we highlight the dynamic modulation of cell-matrix interactions during development. We propose that individual cell-matrix adhesion proteins are best considered as complex proteins that can play multiple, often seemingly contradictory roles, depending upon the context of the microenvironment. In addition, cell-matrix proteins can also exert different short versus long term effects. It is thus important to consider cell behavior in light of the microenvironment because of the constant and dynamic reciprocal interactions occurring between them. Finally, we suggest that analysis of cell-matrix interactions at multiple levels (molecules, cells, tissues) in vivo is critical for an integrated understanding because different information can be acquired from all size scales. PMID:20108219

  3. Extracellular matrix interacts with interferon {alpha} protein: Retention and display of cytotoxicity

    SciTech Connect

    Yoshida, Kimiko; Kondoh, Atsushi; Narumi, Kenta; Yoshida, Teruhiko; Aoki, Kazunori

    2008-11-14

    We have been investigating the efficacy of an intratumoral interferon (IFN)-{alpha} gene transfer against solid cancers, and found that when the gene is transduced into the subcutaneous tumors, IFN-{alpha} concentration is markedly increased in the injected tumor but not in the serum. To explain this effective confinement of IFN-{alpha} to target tissues, we hypothesized that the extracellular matrix in the tumors interacts with IFN-{alpha}. In this study, a solid-phase-binding assay and immunoprecipitation demonstrated that the IFN-{alpha} binds directly to matrix proteins. Immunohistochemical staining showed a co-localization of IFN-{alpha} with pericellular fibronectin. In addition, matrix-bound IFN-{alpha} protein transduced intracellular signaling and potentiated its cytotoxic activity, suggesting that the retention of IFN-{alpha} protein on extracellular matrix is likely to play a role in its in vivo biological activity. The data suggest a therapeutic advantage of the intratumoral IFN-{alpha} gene transfer over the conventional parenteral therapy both in the safety and efficacy.

  4. Analysis of the Aspergillus fumigatus Biofilm Extracellular Matrix by Solid-State Nuclear Magnetic Resonance Spectroscopy

    PubMed Central

    Reichhardt, Courtney; Ferreira, Jose A. G.; Joubert, Lydia-Marie; Clemons, Karl V.; Stevens, David A.

    2015-01-01

    Aspergillus fumigatus is commonly responsible for lethal fungal infections among immunosuppressed individuals. A. fumigatus forms biofilm communities that are of increasing biomedical interest due to the association of biofilms with chronic infections and their increased resistance to antifungal agents and host immune factors. Understanding the composition of microbial biofilms and the extracellular matrix is important to understanding function and, ultimately, to developing strategies to inhibit biofilm formation. We implemented a solid-state nuclear magnetic resonance (NMR) approach to define compositional parameters of the A. fumigatus extracellular matrix (ECM) when biofilms are formed in RPMI 1640 nutrient medium. Whole biofilm and isolated matrix networks were also characterized by electron microscopy, and matrix proteins were identified through protein gel analysis. The 13C NMR results defined and quantified the carbon contributions in the insoluble ECM, including carbonyls, aromatic carbons, polysaccharide carbons (anomeric and nonanomerics), aliphatics, etc. Additional 15N and 31P NMR spectra permitted more specific annotation of the carbon pools according to C-N and C-P couplings. Together these data show that the A. fumigatus ECM produced under these growth conditions contains approximately 40% protein, 43% polysaccharide, 3% aromatic-containing components, and up to 14% lipid. These fundamental chemical parameters are needed to consider the relationships between composition and function in the A. fumigatus ECM and will enable future comparisons with other organisms and with A. fumigatus grown under alternate conditions. PMID:26163318

  5. Novel human-derived extracellular matrix induces in vitro and in vivo vascularization and inhibits fibrosis

    PubMed Central

    Moore, Marc C; Pandolfi, Vittoria; McFetridge, Peter S

    2015-01-01

    The inability to vascularize engineered organs and revascularize areas of infarction has been a major roadblock to delivering successful regenerative medicine therapies to the clinic. These investigations detail an isolated human extracellular matrix derived from the placenta (hPM) that induces vasculogenesis in vitro and angiogenesis in vivo within bioengineered tissues, with significant immune reductive properties. Compositional analysis showed ECM components (fibrinogen, laminin), angiogenic cytokines (angiogenin, FGF), and immune-related cytokines (annexins, DEFA1) in near physiological ratios. Gene expression profiles of endothelial cells seeded onto the matrix displayed upregulation of angiogenic genes (TGFB1, VEGFA), remodeling genes (MMP9, LAMA5) and vascular development genes (HAND2, LECT1). Angiogenic networks displayed a time dependent stability in comparison to current in vitro approaches that degrade rapidly. In vivo, matrix-dosed bioscaffolds showed enhanced angiogenesis and significantly reduced fibrosis in comparison to current angiogenic biomaterials. Implementation of this human placenta derived extracellular matrix provides an alternative to Matrigel and, due to its human derivation, its development may have significant clinical applications leading to advances in therapeutic angiogenesis techniques and tissue engineering. PMID:25725553

  6. Matrix Metalloproteinase 12-Deficiency Augments Extracellular Matrix Degrading Metalloproteinases and Attenuates IL-13–Dependent Fibrosis

    PubMed Central

    Madala, Satish K.; Pesce, John T.; Ramalingam, Thirumalai R.; Wilson, Mark S.; Minnicozzi, Samantha; Cheever, Allen W.; Thompson, Robert W.; Mentink-Kane, Margaret M.; Wynn, Thomas A.

    2011-01-01

    Infection with the parasitic helminth Schistosoma mansoni causes significant liver fibrosis and extracellular matrix (ECM) remodeling. Matrix metalloproteinases (MMP) are important regulators of the ECM by regulating cellular inflammation, extracellular matrix deposition, and tissue reorganization. MMP12 is a macrophage-secreted elastase that is highly induced in the liver and lung in response to S. mansoni eggs, confirmed by both DNA microarray and real-time PCR analysis. However, the function of MMP12 in chronic helminth-induced inflammation and fibrosis is unclear. In this study, we reveal that MMP12 acts as a potent inducer of inflammation and fibrosis after infection with the helminth parasite S. mansoni. Surprisingly, the reduction in liver and lung fibrosis in MMP12-deficient mice was not associated with significant changes in cytokine, chemokine, TGF-β1, or tissue inhibitors of matrix metalloproteinase expression. Instead, we observed marked increases in MMP2 and MMP13 expression, suggesting that Mmp12 was promoting fibrosis by limiting the expression of specific ECM-degrading MMPs. Interestingly, like MMP12, MMP13 expression was highly dependent on IL-13 and type II–IL-4 receptor signaling. However, in contrast to MMP12, expression of MMP13 was significantly suppressed by the endogenous IL-13 decoy receptor, IL-13Rα2. In the absence of MMP12, expression of IL-13Rα2 was significantly reduced, providing a possible explanation for the increased IL-13-driven MMP13 activity and reduced fibrosis. As such, these data suggest important counter-regulatory roles between MMP12 and ECM-degrading enzymes like MMP2, MMP9, and MMP13 in Th2 cytokine-driven fibrosis. PMID:20181883

  7. Nell1-deficient mice have reduced expression of extracellular matrix proteins causing cranial and vertebral defects

    SciTech Connect

    Desai, Jayashree; Shannon, Mark E.; Johnson, Mahlon D.; Ruff, David W.; Hughes, Lori A; Kerley, Marilyn K; Carpenter, D A; Johnson, Dabney K; Rinchik, Eugene M.; Culiat, Cymbeline T

    2006-01-01

    The mammalian Nell1 gene encodes a protein kinase C-b1 (PKC-b1) binding protein that belongs to a new class of cell-signaling molecules controlling cell growth and differentiation. Over-expression of Nell1 in the developing cranial sutures in both human and mouse induces craniosynostosis, the premature fusion of the growing cranial bone fronts. Here, we report the generation, positional cloning and characterization of Nell16R, a recessive, neonatal-lethal point mutation in the mouse Nell1 gene, induced by N-ethyl-N-nitrosourea. Nell16R has a T!A base change that converts a codon for cysteine into a premature stop codon [Cys(502)Ter], resulting in severe truncation of the predicted protein product and marked reduction in steady-state levels of the transcript. In addition to the expected alteration of cranial morphology, Nell16R mutants manifest skeletal defects in the vertebral column and ribcage, revealing a hitherto undefined role for Nell1 in signal transduction in endochondral ossification. Real-time quantitative reverse transcription-PCR assays of 219 genes showed an association between the loss of Nell1 function and reduced expression of genes for extracellular matrix (ECM) proteins critical for chondrogenesis and osteogenesis. Several affected genes are involved in the human cartilage disorder Ehlers-Danlos Syndrome and other disorders associated with spinal curvature anomalies. Nell16R mutant mice are a new tool for elucidating basic mechanisms in osteoblast and chrondrocyte differentiation in the developing skull and vertebral column and understanding how perturbations in the production of ECM proteins can lead to anomalies in these structures.

  8. Periodontal Biological Events Associated with Orthodontic Tooth Movement: The Biomechanics of the Cytoskeleton and the Extracellular Matrix

    PubMed Central

    Feller, L.; Khammissa, R. A. G.; Schechter, I.; Moodley, A.; Thomadakis, G.; Lemmer, J.

    2015-01-01

    The mechanical stimuli generated by orthodontic forces cause deformation of extracellular matrices and cells, vascular changes, inflammation, and the release of active biological agents generating a complex multifactorial sequence of biological events culminating in bone remodelling enabling orthodontic tooth movement. Orthodontic forces on the teeth generate stresses in periodontal tissues according to a number of variables including the type (continuous, interrupted, or intermittent), magnitude, direction, and frequency of the applied load. Whether the strain is compressive or tensile determines whether bone deposition or bone resorption will occur. The mechanically induced strains mediate structural changes in extracellular matrices and in cells, consequently affecting cellular gene expression and function. In the extracellular matrix, mechanosensing molecules integrated into the structure of various proteins can be activated upon load-induced protein unfolding. These specialized molecules have the capacity to sense and then to convert microenvironmental biomechanical stimuli into intracellular biochemical signals that interact to generate a coordinated tissue response. It is also possible that the applied force may directly cause nuclear deformation with configurational changes in chromatin, thus influencing gene expression. In this review article we summarize the current general concepts of mechanotransduction influencing the remodelling of periodontal tissues thus enabling tooth movement in response to applied orthodontic loads. PMID:26351659

  9. SMAD4 mutations causing Myhre syndrome result in disorganization of extracellular matrix improved by losartan

    PubMed Central

    Piccolo, Pasquale; Mithbaokar, Pratibha; Sabatino, Valeria; Tolmie, John; Melis, Daniela; Schiaffino, Maria Cristina; Filocamo, Mirella; Andria, Generoso; Brunetti-Pierri, Nicola

    2014-01-01

    Myhre syndrome (MS, MIM 139210) is a connective tissue disorder that presents with short stature, short hands and feet, facial dysmorphic features, muscle hypertrophy, thickened skin, and deafness. Recurrent missense mutations in SMAD4 encoding for a transducer mediating transforming growth factor β (TGF-β) signaling are responsible for MS. We found that MS fibroblasts showed increased SMAD4 protein levels, impaired matrix deposition, and altered expression of genes encoding matrix metalloproteinases and related inhibitors. Increased TGF-β signaling and progression of aortic root dilation in Marfan syndrome can be prevented by the antihypertensive drug losartan, a TGF-β antagonists and angiotensin-II type 1 receptor blocker. Herein, we showed that losartan normalizes metalloproteinase and related inhibitor transcript levels and corrects the extracellular matrix deposition defect in fibroblasts from MS patients. The results of this study may pave the way toward therapeutic applications of losartan in MS. PMID:24398790

  10. Age-related macular degeneration and changes in the extracellular matrix

    PubMed Central

    Nita, Małgorzata; Strzałka-Mrozik, Barbara; Grzybowski, Andrzej; Mazurek, Urszula; Romaniuk, Wanda

    2014-01-01

    Age-related macular degeneration (AMD) is the leading cause of permanent, irreversible, central blindness (scotoma in the central visual field that makes reading and writing impossible, stereoscopic vision, recognition of colors and details) in patients over the age of 50 years in European and North America countries, and an important role is attributed to disorders in the regulation of the extracellular matrix (ECM). The main aim of this article is to present the crucial processes that occur on the level of Bruch’s membrane, with special consideration of the metalloproteinase substrates, metalloproteinase, and tissue inhibitor of metalloproteinase (TIMP). A comprehensive review of the literature was performed through MEDLINE and PubMed searches, covering the years 2005–2012, using the following keywords: AMD, extracellular matrix, metalloproteinases, tissue inhibitors of metalloproteinases, Bruch’s membrane, collagen, elastin. In the pathogenesis of AMD, a significant role is played by collagen type I and type IV; elastin; fibulin-3, -5, and -6; matrix metalloproteinase (MMP)-2, MMP-9, MMP-14, and MMP-1; and TIMP-3. Other important mechanisms include: ARMS2 and HTR1 proteins, the complement system, the urokinase plasminogen activator system, and pro-renin receptor activation. Continuous rebuilding of the extracellular matrix occurs in both early and advanced AMD, simultaneously with the dysfunction of retinal pigment epithelium (RPE) cells and endothelial cells. The pathological degradation or accumulation of ECM structural components are caused by impairment or hyperactivity of specific MMPs/TIMPs complexes, and is also endangered by the influence of other mechanisms connected with both genetic and environmental factors. PMID:24938626

  11. Extracellular matrix is a source of mitogenically active platelet-derived growth factor.

    PubMed

    Field, S L; Khachigian, L M; Sleigh, M J; Yang, G; Vandermark, S E; Hogg, P J; Chesterman, C N

    1996-08-01

    Platelet-derived growth factor (PDGF) is a chemotactic and mitogenic agent for fibroblasts and smooth muscle cells and plays a key role in the development of atherosclerotic lesions. PDGF is produced by a number of normal and transformed cell types and occurs as homo- or heterodimers of A and B polypeptide chains. Using Chinese hamster ovary (CHO) cells transfected with various forms of PDGF, we have previously shown that PDGF A(s) (short splice version) is secreted, PDGF A(l) (long splice version) predominantly extracellular matrix-associated, and PDGF B divided between medium, cells, and matrix. In the present study we have demonstrated the mitogenic activity of matrix-localized PDGF in artificial and more physiologically relevant models by culturing Balb/c-3T3 cells (3T3), human foreskin fibroblasts (HFF), and rabbit aortic smooth muscle cells (SMC) on extracellular matrix (ECM) laid down by PDGF-expressing CHO cells and human umbilical vein endothelial cells (HUVEC). These cells responded to the local growth stimulus of PDGF-containing CHO ECM and HUVEC ECM. We showed that 3T3 cells required proteolytic activity to utilize matrix-localized PDGF, as aprotinin and epsilon-ACA inhibited growth and 3T3 cells were shown to possess plasminogen activator activity. HFF and SMC did not appear to require proteolytic activity (including metalloproteinase and serine protease activity) as a prerequisite for mitogenesis but were able to access immobilized PDGF by contact with the matrix. An understanding of the mechanisms whereby the utilization of stored PDGF is controlled in situations of excessive cellular proliferation will aid in the development of therapy for these conditions. PMID:8707868

  12. Decellularization of porcine skeletal muscle extracellular matrix for the formulation of a matrix hydrogel: a preliminary study.

    PubMed

    Fu, Yuehe; Fan, Xuejiao; Tian, Chunxiang; Luo, Jingcong; Zhang, Yi; Deng, Li; Qin, Tingwu; Lv, Qing

    2016-04-01

    Extracellular matrix (ECM) hydrogels are used as scaffolds to facilitate the repair and reconstruction of tissues. This study aimed to optimize the decellularization process of porcine skeletal muscle ECM and to formulate a matrix hydrogel scaffold. Five multi-step methods (methods A-E) were used to generate acellular ECM from porcine skeletal muscle [rinsing in SDS, trypsin, ethylenediaminetetraacetic acid (EDTA), Triton X-100 and/or sodium deoxycholate at 4-37°C]. The resulting ECM was evaluated using haematoxylin and eosin, 4-6-diamidino-2-phenylindole (DAPI) staining, and DNA quantification. Acellular matrix was dissolved in pepsin and gelled at 37°C. Hydrogel response to temperature was observed in vivo and in vitro. ECM components were assessed by Masson, Sirius red, and alcian blue staining, and total protein content. Acellular porcine skeletal muscle exhibited a uniform translucent white appearance. No intact nuclear residue was detected by haematoxylin and eosin staining, while DAPI staining showed a few nuclei in the matrixes produced by methods B, C, and D. Method A generated a gel that was too thin for gelation. However, the matrix obtained by rinsing in 0.2% trypsin/0.1% EDTA, 0.5% Triton X-100, and 1% Triton X-100/0.2% sodium deoxycholate was nuclei-free and produced a viscous solution that formed a structurally stable white jelly-like hydrogel. The residual DNA content of this solution was 49.37 ± 0.72 ng/mg, significantly less than in fresh skeletal muscle, and decreased to 19.22 ± 0.85 ng/mg after gelation (P < 0.05). The acellular matrix was rich in collagen and glycosaminoglycan, with a total protein concentration of 64.8 ± 6.9%. An acellular ECM hydrogel from porcine skeletal muscle was efficiently produced. PMID:26781342

  13. Tendon extracellular matrix damage, degradation and inflammation in response to in vitro overload exercise

    PubMed Central

    Thorpe, Chavaunne T.; Chaudhry, Saira; Riley, Graham P.; Birch, Helen L.; Clegg, Peter D.; Screen, Hazel R.C.

    2015-01-01

    ABSTRACT The role of inflammation in tendon injury is uncertain and a topic of current interest. In vitro studies of tendon accelerated overload damage can serve as a valuable source of information on the early stages of tendinopathy. Viable fascicle bundles from bovine flexor tendons were subjected to cyclic uniaxial loading from 1–10% strain. Immuno‐staining for inflammatory markers and matrix degradation markers was performed on the samples after mechanical testing. Loaded samples exhibited visible extracellular matrix damage, with disrupted collagen fibers and fiber kinks, and notable damage to the interfascicular matrix. Inflammatory markers COX‐2 and IL‐6 were only expressed in the cyclically loaded samples. Collagen degradation markers MMP‐1 and C1,2C were colocalized in many areas, with staining occurring in the interfascicular matrix or the fascicular tenocytes. These markers were present in control samples, but staining became increasingly intense with loading. Little MMP‐3 or MMP‐13 was evident in control sections. In loaded samples, some sections showed intense staining of these markers, again localized to interfascicular regions. This study suggests that inflammatory markers may be expressed rapidly after tendon overload exercise. Interestingly, both inflammation and damage‐induced matrix remodeling seem to be concentrated in, or in the vicinity of, the highly cellular interfascicular matrix. © 2015 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. J Orthop Res 33:889–897, 2015. PMID:25721513

  14. Downregulation of the Yes-Associated Protein Is Associated with Extracellular Matrix Disorders in Ascending Aortic Aneurysms

    PubMed Central

    Li, Haiyang; Jiang, Wenjian; Ren, Weihong; Guo, Dong; Guo, Jialong; Wang, Xiaolong; Liu, Yuyong; Lan, Feng; Du, Jie; Zhang, Hongjia

    2016-01-01

    Previous studies indicate that extracellular matrix (ECM) disorders lead to the apoptosis of Vascular Smooth Muscle Cells (VSMCs), which impairs the aortic wall by reducing the generation of elastic fibers, and ultimately result in ascending aortic aneurysm. The critical role of the Yes-associated protein (YAP) has been elucidated in cardiac/SMC proliferation during cardiovascular development. However, the association of YAP expression and extracellular matrix disorders in ascending aortic aneurysms is not clear. Here, we present for the first time that the downregulation of YAP in VSMCs is associated with ECM disorders of the media in ascending aortic aneurysms. We found that aortic ECM deteriorated with increased apoptotic VSMCs. Moreover, expression of YAP was dramatically reduced in the aortic walls of patients with ascending aortic aneurysms, while the normal aortic samples exhibited abundant YAP in the VSMCs. These results suggest that downregulation of YAP leads to apoptosis of VSMCs, which are essential for the homeostasis of the aortic wall. The resultant ECM disorders affect aortic structure and function and contribute to the development of ascending aortic aneurysms. In summary, through assessment of clinical samples, we revealed the association between downregulation of YAP in VSMCs and the development of ascending aortic aneurysms, providing new insight into the pathogenesis of this disease. PMID:26904131

  15. [Extracellular matrix remodeling myocardium of the left ventricle rats with experimental heart failure after perindopril and melatonin administration].

    PubMed

    Liskova, Iu V; Salikova, S P; Stadnikov, A A

    2014-01-01

    The aim of the study was to examine the reorganization of extracellular matrix left ventricular myocardium (LVM) rats (n = 38) with experimental heart failure (EHF) as affected by perindopril and melatonin administration. Rats LVM was studied using the methods of light microscopy, immunocytochemistry and morphometry. At day 14 of EHF, marked mosaic staining cardiomyocytes acid dyes, manifest hemodynamic disturbances: venous and capillary congestion, lymphostasis, perivascular and interstitial edema, there was a increase the volume density (OD) stroma, high activity of MMP-1 and TIMP-1. After modeling the EHF to 28 days in LVM, a further increase of hemodynamic met, cardiosclerosis areas and perivascular sclerosis, a significant increase OD stroma, observed decrease in the expression of MMP-1 and TIMP-1. In rats given perindopril and melatonin for 14 days, there was a regression of pathological changes in the balance was maintained MMP-1/TIMP-1 ratio close to the group of intact rats. Cardioprotective effects of perindopril and melatonin on extracellular matrix LVM of rats with EHF is discussed. PMID:25702403

  16. MicroRNA-375 Suppresses Extracellular Matrix Degradation and Invadopodial Activity in Head and Neck Squamous Cell Carcinoma

    PubMed Central

    Jimenez, Lizandra; Sharma, Ved P.; Condeelis, John; Harris, Thomas; Ow, Thomas J.; Prystowsky, Michael B.; Childs, Geoffrey; Segall, Jeffrey E.

    2015-01-01

    Context Head and neck squamous cell carcinoma (HNSCC) is a highly invasive cancer with an association with locoregional recurrence and lymph node metastasis. We have previously reported that low microRNA-375 (miR-375) expression levels correlate with poor patient survival, increased locoregional recurrence, and distant metastasis. Increasing miR-375 expression in HNSCC cell lines to levels found in normal cells results in suppressed invasive properties. HNSCC invasion is mediated in part by invadopodia-associated degradation of the extracellular matrix. Objective To determine whether elevated miR-375 expression in HNSCC cell lines also affects invadopodia formation and activity. Design For evaluation of the matrix degradation properties of the HNSCC lines, an invadopodial matrix degradation assay was used. The total protein levels of invadopodia-associated proteins were measured by Western blot analyses. Immunoprecipitation experiments were conducted to evaluate the tyrosine phosphorylation state of cortactin. Human Protease Arrays were used for the detection of the secreted proteases. Quantitative real time–polymerase chain reaction measurements were used to evaluate the messenger RNA (mRNA) expression of the commonly regulated proteases. Results Increased miR-375 expression in HNSCC cells suppresses extracellular matrix degradation and reduces the number of mature invadopodia. Higher miR-375 expression does not reduce cellular levels of selected invadopodia-associated proteins, nor is tyrosine phosphorylation of cortactin altered. However, HNSCC cells with higher miR-375 expression had significant reductions in the mRNA expression levels and secreted levels of specific proteases. Conclusions MicroRNA-375 regulates invadopodia maturation and function potentially by suppressing the expression and secretion of proteases. PMID:26172508

  17. Cytoskeletal filament assembly and the control of cell spreading and function by extracellular matrix

    NASA Technical Reports Server (NTRS)

    Mooney, D. J.; Langer, R.; Ingber, D. E.

    1995-01-01

    This study was undertaken to analyze how cell binding to extracellular matrix produces changes in cell shape. We focused on the initial process of cell spreading that follows cell attachment to matrix and, thus, cell 'shape' changes are defined here in terms of alterations in projected cell areas, as determined by computerized image analysis. Cell spreading kinetics and changes in microtubule and actin microfilament mass were simultaneously quantitated in hepatocytes plated on different extracellular matrix substrata. The initial rate of cell spreading was highly dependent on the matrix coating density and decreased from 740 microns 2/h to 50 microns 2/h as the coating density was lowered from 1000 to 1 ng/cm2. At approximately 4 to 6 hours after plating, this initial rapid spreading rate slowed and became independent of the matrix density regardless of whether laminin, fibronectin, type I collagen or type IV collagen was used for cell attachment. Analysis of F-actin mass revealed that cell adhesion to extracellular matrix resulted in a 20-fold increase in polymerized actin within 30 minutes after plating, before any significant change in cell shape was observed. This was followed by a phase of actin microfilament disassembly which correlated with the most rapid phase of cell extension and ended at about 6 hours; F-actin mass remained relatively constant during the slow matrix-independent spreading phase. Microtubule mass increased more slowly in spreading cells, peaking at 4 hours, the time at which the transition between rapid and slow spreading rates was observed. However, inhibition of this early rise in microtubule mass using either nocodazole or cycloheximide did not prevent this transition. Use of cytochalasin D revealed that microfilament integrity was absolutely required for hepatocyte spreading whereas interference with microtubule assembly (using nocodazole or taxol) or protein synthesis (using cycloheximide) only partially suppressed cell extension. In

  18. Interactions between Skeletal Muscle Myoblasts and their Extracellular Matrix Revealed by a Serum Free Culture System

    PubMed Central

    Chaturvedi, Vishal; Dye, Danielle E.; Kinnear, Beverley F.; van Kuppevelt, Toin H.; Grounds, Miranda D.; Coombe, Deirdre R.

    2015-01-01

    Decellularisation of skeletal muscle provides a system to study the interactions of myoblasts with muscle extracellular matrix (ECM). This study describes the efficient decellularisation of quadriceps muscle with the retention of matrix components and the use of this matrix for myoblast proliferation and differentiation under serum free culture conditions. Three decellularisation approaches were examined; the most effective was phospholipase A2 treatment, which removed cellular material while maximizing the retention of ECM components. Decellularised muscle matrices were then solubilized and used as substrates for C2C12 mouse myoblast serum free cultures. The muscle matrix supported myoblast proliferation and differentiation equally as well as collagen and fibronectin. Immunofluorescence analyses revealed that myoblasts seeded on muscle matrix and fibronectin differentiated to form long, well-aligned myotubes, while myoblasts seeded on collagen were less organized. qPCR analyses showed a time dependent increase in genes involved in skeletal muscle differentiation and suggested that muscle-derived matrix may stimulate an increased rate of differentiation compared to collagen and fibronectin. Decellularized whole muscle three-dimensional scaffolds also supported cell adhesion and spreading, with myoblasts aligning along specific tracts of matrix proteins within the scaffolds. Thus, under serum free conditions, intact acellular muscle matrices provided cues to direct myoblast adhesion and migration. In addition, myoblasts were shown to rapidly secrete and organise their own matrix glycoproteins to create a localized ECM microenvironment. This serum free culture system has revealed that the correct muscle ECM facilitates more rapid cell organisation and differentiation than single matrix glycoprotein substrates. PMID:26030912

  19. The dynamic sclera: extracellular matrix remodeling in normal ocular growth and myopia development.

    PubMed

    Harper, Angelica R; Summers, Jody A

    2015-04-01

    Myopia is a common ocular condition, characterized by excessive elongation of the ocular globe. The prevalence of myopia continues to increase, particularly among highly educated groups, now exceeding 80% in some groups. In parallel with the increased prevalence of myopia, are increases in associated blinding ocular conditions including glaucoma, retinal detachment and macular degeneration, making myopia a significant global health concern. The elongation of the eye is closely related to the biomechanical properties of the sclera, which in turn are largely dependent on the composition of the scleral extracellular matrix. Therefore an understanding of the cellular and extracellular events involved in the regulation of scleral growth and remodeling during childhood and young adulthood will provide future avenues for the treatment of myopia and its associated ocular complications. PMID:25819458

  20. Extracellular matrix and cell shape: potential control points for inhibition of angiogenesis

    NASA Technical Reports Server (NTRS)

    Ingber, D.

    1991-01-01

    Capillary endothelial (CE) cells require two extracellular signals in order to switch from quiescence to growth and back to differentiation during angiogenesis: soluble angiogenic factors and insoluble extracellular matrix (ECM) molecules. Soluble endothelial mitogens, such as basic fibroblast growth factor (FGF), act over large distances to trigger capillary growth, whereas ECM molecules act locally to modulate cell responsiveness to these soluble cues. Recent studies reveal that ECM molecules regulate CE cell growth and differentiation by modulating cell shape and by activating intracellular chemical signaling pathways inside the cell. Recognition of the importance of ECM and cell shape during capillary morphogenesis has led to the identification of a series of new angiogenesis inhibitors. Elucidation of the molecular mechanism of capillary regulation may result in development of even more potent angiogenesis modulators in the future.

  1. Semicarbazide-sensitive amine oxidase and extracellular matrix deposition by smooth-muscle cells

    NASA Technical Reports Server (NTRS)

    Langford, Shannon D.; Trent, Margaret B.; Boor, Paul J.

    2002-01-01

    We have recently reported in vivo disruption of collagen and elastin architecture within blood vessel walls resulting from the selective inhibition of the enzyme semicarbazide-sensitive amine oxidase (SSAO). This study further investigates the effects of SSAO inhibition on extracellular matrix deposition by smooth-muscle cells (SMCs) cultured from neonatal rat hearts. SMCs were characterized, SSAO activity was measured, and soluble and insoluble collagen and elastin in the extracellular matrix (ECM) were quantified. Cultured neonatal rat heart SMC exhibited a monotypic synthetic phenotype that likely represents a myofibroblast. Detectable levels of SSAO activity present throughout 30-d culture peaked at 7-14 d, coinciding with the production of ECM. The addition of enzyme inhibitors and alternate SSAO substrates (benzylamine) produced varied and, in some cases, marked changes in SSAO activity as well as in the composition of mature and soluble matrix components. Similar to our previous in vivo findings, in vitro SSAO inhibition produced aberrations in collagen and elastin deposition by heart SMC. Because changes in SSAO activity are associated with cardiovascular pathologic states, this enzyme may play a protective or modulating role by regulating ECM production during pathologic insult.

  2. Lipoprotein lipase enhances binding of lipoproteins to heparan sulfate on cell surfaces and extracellular matrix.

    PubMed Central

    Eisenberg, S; Sehayek, E; Olivecrona, T; Vlodavsky, I

    1992-01-01

    Lipoprotein lipase enhances binding at 4 degrees C of human plasma lipoproteins (chylomicrons, VLDL, intermediate density lipoprotein, LDL, and HDL3) to cultured fibroblasts and hepG-2 cells and to extracellular matrix. Heparinase treatment of cells and matrix reduces the lipoprotein lipase enhanced binding by 90-95%. Lipoprotein lipase causes only a minimal effect on the binding of lipoproteins to heparan sulfate deficient mutant Chinese hamster ovary cells while it promotes binding to wild type cells that is abolished after heparinase treatment. With 125I-LDL, lipoprotein lipase also enhances uptake and proteolytic degradation at 37 degrees C by normal human skin fibroblasts but has no effect in heparinase-treated normal cells or in LDL receptor-negative fibroblasts. These observations prove that lipoprotein lipase causes, predominantly, binding of lipoproteins to heparan sulfate at cell surfaces and in extracellular matrix rather than to receptors. This interaction brings the lipoproteins into close proximity with cell surfaces and may promote metabolic events that occur at the cell surface, including facilitated transfer to cellular receptors. Images PMID:1430223

  3. Selective enrichment of microRNAs in extracellular matrix vesicles produced by growth plate chondrocytes.

    PubMed

    Lin, Zhao; Rodriguez, Nicholas E; Zhao, Junjun; Ramey, Allison N; Hyzy, Sharon L; Boyan, Barbara D; Schwartz, Zvi

    2016-07-01

    Matrix vesicles (MVs) are membrane organelles found in the extracellular matrix of calcifying cells, which contain matrix processing enzymes and regulate the extracellular environment via action of these enzymes. It is unknown whether MVs are also exosomic mediators of cell-cell communication via transfer of RNA material, and specifically, microRNA (miRNA). We investigated the presence of RNA in MVs isolated from cultures of costochondral growth zone chondrocytes. Our results showed that the average yield of MV RNA was 1.93±0.78ng RNA/10(4) cells, which was approximately 0.1% of the parent cell's total RNA. MV RNA was well-protected from RNase by the lipid membrane and was highly enriched in small RNA molecules compared to cells. Moreover, coding and non-coding small RNAs in MVs were in proportions that differed from parent cells. Enrichment of specific miRNAs was consistently observed in all three miRNA detection platforms that we used, suggesting that miRNAs are selectively packaged into MVs. MV-enriched miRNAs were related to different signaling pathways associated with bone formation. This study suggests a significant role for MVs as "matrisomes" in cell-cell communication in cartilage and bone development via transfer of specific miRNAs. PMID:27080510

  4. Transglutaminase 2 interactions with extracellular matrix proteins as probed with celiac disease autoantibodies.

    PubMed

    Cardoso, Inês; Stamnaes, Jorunn; Andersen, Jan Terje; Melino, Gerry; Iversen, Rasmus; Sollid, Ludvig M

    2015-06-01

    Transglutaminases have been implicated in various human diseases. A prominent example is the involvement of transglutaminase 2 (TG2) in the gluten-sensitive enteropathy celiac disease, where the enzyme is both the target of autoantibodies and responsible for the generation of immunogenic gluten epitopes. Here, we aimed to characterize the microenvironment of TG2 in the extracellular matrix (ECM) in order to gain insights into the antigenic structures that are recognized by autoantibodies in celiac disease. A panel of TG2-specific mAbs established from gut plasma cells of celiac disease patients was employed as probes to characterize the interactions between TG2 and ECM constituents. With immunofluorescence staining, microplate protein-binding and surface plasmon resonance assays, we found that the main epitope (epitope 1) recognized by TG2-specific gut plasma cells overlaps with the fibronectin (FN)-binding site of TG2. Furthermore, we found that the same TG2 amino acids that are involved in binding of epitope 1 mAbs are also important for efficient binding of FN. Notably, epitope 1 mAbs recognize TG2 in tissue sections, suggesting that some TG2 in the extracellular matrix has interaction partners in addition to FN. We demonstrate that collagen VI is a strong candidate, on the basis of its tissue expression pattern and ability to bind TG2. Collagen VI may thus serve as a matrix for deposition of TG2 in a context that can also be recognized by epitope 1-targeting autoantibodies. PMID:25808416

  5. Platelets and plasma stimulate sheep rotator cuff tendon tenocytes when cultured in an extracellular matrix scaffold.

    PubMed

    Kelly, Brian A; Proffen, Benedikt L; Haslauer, Carla M; Murray, Martha M

    2016-04-01

    The addition of platelet-rich plasma (PRP) to rotator cuff repair has not translated into improved outcomes after surgery. However, recent work stimulating ligament healing has demonstrated improved outcomes when PRP or whole blood is combined with an extracellular matrix carrier. The objective of this study was to evaluate the effect of three components of blood (plasma, platelets, and macrophages) on the in vitro activity of ovine rotator cuff cells cultured in an extracellular matrix environment. Tenocytes were obtained from six ovine infraspinatus tendons and cultured over 14 days in an extracellular matrix scaffold with the following additives: (1) plasma (PPP), (2) plasma and platelets (PAP), (3) plasma and macrophages (PPPM), (4) plasma, platelets and macrophages (PAPM), (5) phosphate buffered saline (PBS), and (6) PBS with macrophages (PBSM). Assays measuring cellular metabolism (AlamarBlue), proliferation (Quantitative DNA assay), synthesis of collagen and cytokines (SIRCOL, TNF-α and IL-10 ELISA, and MMP assay), and collagen gene expression (qPCR) were performed over the duration of the experiment, as well as histology at the conclusion. Plasma was found to stimulate cell attachment and spreading on the scaffold, as well as cellular proliferation. Platelets also stimulated cell proliferation, cellular metabolism, transition of cells to a myofibroblast phenotype, and contraction of the scaffolds. The addition of macrophages did not have any significant effect on the sheep rotator cuff cells in vitro. In vivo studies are needed to determine whether these changes in cellular function will translate into improved tendon healing. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:623-629, 2016. PMID:26419602

  6. The isolation of cell derived extracellular matrix constructs using sacrificial open-cell foams.

    PubMed

    Wolchok, Jeffrey C; Tresco, Patrick A

    2010-12-01

    Extracellular matrix derived from human and animal tissues is being used to repair and reconstruct a variety of tissues clinically. The utility of such constructs is limited by the geometry, composition and constitutive properties of the tissue or organ from which the ECM is harvested. To address this limitation, we have developed an approach to isolate extracellular matrix in bulk from populations of living cells grown in culture on three-dimensional substrates. Human biopsy derived fibroblasts were seeded within open-cell foams and cultured in-vitro for periods up to three weeks, after which the synthetic component was removed by incubation in a water miscible solvent. After several wash steps and lyophilization, a white, lacy, multi-molecular construct was isolated. Tandem mass spectroscopy showed that it contained 22 extracellular matrix constituents, including such proteins and proteoglycans as collagen type I and type III, fibronectin, transforming growth factor beta, decorin and biglycan among others. On average 47 mg of construct was isolated for each gram of synthetic substrate initially seeded with cells. The biomaterial harvested from human tracheal fibroblasts had an elastic modulus (250 kPa) and a composition similar to that of human vocal fold tissue, and supported reseeding with human tracheal derived fibroblasts. An important finding was that the approach was useful in isolating ECM from a variety of cell lineages and developmental stages including skin fibroblasts, brain derived astrocytes and mesenchymal stem cells. The results, together with the archival literature, suggest that the approach can be used to produce a range of cell derived constructs with unique physical and chemical attributes for a variety of research and medical applications. PMID:20950855

  7. The contribution of the extracellular matrix to gravisensing in characean cells

    NASA Technical Reports Server (NTRS)

    Wayne, R.; Staves, M. P.; Leopold, A. C.

    1992-01-01

    The cell-extracellular matrix junction, which includes the cell wall and the outer surface of the plasma membrane, may be an essential region for the perception of gravity by the internodal cells of Chara corallina. Typically, when an internodal cell is oriented vertically, the downwardly directed cytoplasmic stream travels at a velocity that is 10% faster than that of the upwardly directed stream. However when the cells are treated with impermeant hydrolytic enzymes that partially digest cellulose or hemicellulose, the cells lose their ability to respond to gravity even though streaming continues. By contrast, enzymes that digest pectins have no effect on the gravity-induced polarity of cytoplasmic streaming. Furthermore, gravisensing is sensitive to protease treatment; Proteinase K, thermolysin and collagenase but not trypsin, alpha-chymotrypsin or carboxypeptidase B, inhibit gravisensing. These findings indicate that proteins in the cell-extracellular matrix junction may be required for gravisensing. Moreover, the tetrapeptide Arg-Gly-Asp-Ser (RGDS) inhibits gravisensing in a concentration-dependent manner, indicating that the gravireceptor may be an integrin-like protein. The macromolecules necessary for gravisensing have been localized to the cell ends. As a consequence of the exoplasmic site of action of the enzymes and the tetrapeptides, we interpret the results to mean that they are acting on the gravireceptor, although we cannot eliminate the possibility that they are acting on the signal transduction chain. On the whole, our observations indicate that the cell-extracellular matrix junction is a sine qua non for graviperception in statolith-free Chara internodal cells and we suggest that the gravireceptor is located in this region.

  8. Virus activated filopodia promote human papillomavirus type 31 uptake from the extracellular matrix

    PubMed Central

    Smith, Jessica L.; Lidke, Diane S.; Ozbun, Michelle A.

    2011-01-01

    Human papillomaviruses (HPVs), etiological agents of epithelial tumors and cancers, initiate infection of basal human keratinocytes (HKs) facilitated by wounding. Virions bind to HKs and their secreted extracellular matrix (ECM), but molecular roles for wounding or ECM binding during infection are unclear. Herein we demonstrate HPV31 activates signals promoting cytoskeletal rearrangements and virion transport required for internalization and infection. Activation of tyrosine and PI3 kinases precedes induction of filopodia whereon virions are transported toward the cell body. Coupled with loss of ECM bound virions this supports a model whereby virus activated filopodial transport contributes to increased and protracted virion uptake into susceptible cells. PMID:18834609

  9. The Extracellular Matrix in Epithelial Ovarian Cancer – A Piece of a Puzzle

    PubMed Central

    Cho, Angela; Howell, Viive M.; Colvin, Emily K.

    2015-01-01

    Epithelial ovarian cancer is the fifth leading cause of cancer-related deaths in women and the most lethal gynecological malignancy. Extracellular matrix (ECM) is an integral component of both the normal and tumor microenvironment. ECM composition varies between tissues and is crucial for maintaining normal function and homeostasis. Dysregulation and aberrant deposition or loss of ECM components is implicated in ovarian cancer progression. The mechanisms by which tumor cells induce ECM remodeling to promote a malignant phenotype are yet to be elucidated. A thorough understanding of the role of the ECM in ovarian cancer is needed for the development of effective biomarkers and new therapies. PMID:26579497

  10. Native Cardiac Extracellular Matrix Hydrogels for Cultivation of Human Stem Cell-Derived Cardiomyocytes

    PubMed Central

    Freytes, Donald O; O’Neill, John D; Duan-Arnold, Yi; Wrona, Emily; Vunjak-Novakovic, Gordana

    2015-01-01

    Summary Biomaterial scaffolds made of native and synthetic materials are designed to serve as a structural and informational template for cell attachment and tissue formation. The use of native extracellular matrix (ECM) is of special interest for the culture of cardiac stem and progenitor cells due to the presence of intrinsic regulatory factors regulating cardiac function. We describe here how to obtain native ECM hydrogels from porcine hearts for the culture of human embryonic, induced pluripotent, and somatic stem cells for cardiac tissue engineering and regenerative medicine applications. PMID:25070328

  11. Suppression of ICE and Apoptosis in Mammary Epithelial Cells by Extracellular Matrix

    SciTech Connect

    Boudreau, Nancy; Sympson, C. J.; Werb, Zena; Bissell, Mina J.

    1994-12-01

    Apoptosis (programmed cell death) plays a major role in development and tissue regeneration. Basement membrane extracellular matrix (ECM), but not fibronectin or collagen, was shown to suppress apoptosis of mammary epithelial cells in tissue culture and in vivo. Apoptosis was induced by antibodies to beta 1 integrins or by overexpression of stromelysin-1, which degrades ECM. Expression of interleukin-1 beta converting enzyme (ICE) correlated with the loss of ECM, and inhibitors of ICE activity prevented apoptosis. These results suggest that ECM regulates apoptosis in mammary epithelial cells through an integrin-dependent negative regulation of ICE expression.

  12. Of extracellular matrix, scaffolds, and signaling: Tissuearchitectureregulates development, homeostasis, and cancer

    SciTech Connect

    Nelson, Celeste M.; Bissell, Mina J.

    2006-03-09

    The microenvironment surrounding cells influences gene expression, such that a cell's behavior is largely determined by its interactions with the extracellular matrix, neighboring cells, and soluble cues released locally or by distant tissues. We describe the essential role of context and organ structure in directing mammary gland development and differentiated function, and in determining response to oncogenic insults including mutations. We expand on the concept of 'dynamic reciprocity' to present an integrated view of development, cancer, and aging, and posit that genes are like piano keys: while essential, it is the context that makes the music.

  13. Emerging Implications for Extracellular Matrix-Based Technologies in Vascularized Composite Allotransplantation

    PubMed Central

    Londono, Ricardo; Gorantla, Vijay S.; Badylak, Stephen F.

    2016-01-01

    Despite recent progress in vascularized composite allotransplantation (VCA), limitations including complex, high dose immunosuppression regimens, lifelong risk of toxicity from immunosuppressants, acute and most critically chronic graft rejection, and suboptimal nerve regeneration remain particularly challenging obstacles restricting clinical progress. When properly configured, customized, and implemented, biomaterials derived from the extracellular matrix (ECM) retain bioactive molecules and immunomodulatory properties that can promote stem cell migration, proliferation and differentiation, and constructive functional tissue remodeling. The present paper reviews the emerging implications of ECM-based technologies in VCA, including local immunomodulation, tissue repair, nerve regeneration, minimally invasive graft targeted drug delivery, stem cell transplantation, and other donor graft manipulation. PMID:26839554

  14. Of Extracellular Matrix, Scaffolds, and Signaling: Tissue Architecture Regulates Development, Homeostasis, and Cancer

    PubMed Central

    Nelson, Celeste M.; Bissell, Mina J.

    2010-01-01

    The microenvironment influences gene expression so that the behavior of a cell is largely determined by its interactions with the extracellular matrix, neighboring cells, and soluble local and systemic cues. We describe the essential roles of context and organ structure in directing mammary gland development and differentiated function and in determining the response to oncogenic insults, including mutations. We expand on the concept of “dynamic reciprocity” to present an integrated view of development, cancer, and aging and posit that genes are like the keys on a piano: Although they are essential, it is the context that makes the music. PMID:16824016

  15. Natural cardiac extracellular matrix hydrogels for cultivation of human stem cell-derived cardiomyocytes.

    PubMed

    Freytes, Donald O; O'Neill, John D; Duan-Arnold, Yi; Wrona, Emily A; Vunjak-Novakovic, Gordana

    2014-01-01

    Biomaterial scaffolds made of natural and synthetic materials are designed to serve as a structural and informational template for cell attachment and tissue formation. The use of native extracellular matrix (ECM) is of special interest for the culture of cardiac stem and progenitor cells due to the presence of intrinsic regulatory factors regulating cardiac function. We describe here how to obtain native ECM hydrogels from porcine hearts for the culture of human embryonic, induced pluripotent, and somatic stem cells for cardiac tissue engineering and regenerative medicine applications. PMID:25070328

  16. Overload-Induced Skeletal Muscle Extracellular Matrix Remodeling And Myofiber Growth in Mice Lacking IL-6

    PubMed Central

    White, James P.; Reecy, James M.; Washington, Tyrone A.; Sato, Shuichi; Le, Michael E.; Davis, J. Mark; Wilson, L. Britt; Carson, James A.

    2011-01-01

    Aim Overloading healthy skeletal muscle produces myofiber hypertrophy and extracellular matrix remodeling, and these processes are thought to be interdependent for producing muscle growth. Inflammatory cytokine interleukin-6 (IL-6) gene expression is induced in overloaded skeletal muscle, and the loss of this IL-6 induction can attenuate the hypertrophic response to overload. Although the overload induction of IL-6 in skeletal muscle may be an important regulator of inflammatory processes and satellite cell proliferation, less is known about its role in the regulation of extracellular matrix remodeling. The purpose of the current study was to examine if overload-induced extracellular matrix remodeling, muscle growth, and associated gene expression were altered in mice that lack IL-6, when compared to wild-type mice. Methods Male C57/BL6 (WT) and C57/BL6 × IL-6-/- (IL-6-/-) mice (10 wks of age) were assigned to either a sham control or synergist ablation overload (OV) treatments for 3 or 21 days. Results Plantaris muscle mass increased 59% in WT and 116% in IL-6-/- mice after 21d OV. Myofiber CSA was also increased by 21d OV in both WT and IL-6-/- mice. Overload induced a 2-fold greater increase in the volume of non-contractile tissue in IL-6-/- muscle as compared to WT. Overload also induced a significantly greater accumulation of hydroxyproline and procollagen-1 mRNA in IL-6-/- muscle, when compared to WT muscle after 21d OV. TGF-β and IGF-1 mRNA expression were also induced to a greater extent in IL-6-/- muscle when compared to WT muscle after 21d OV. There was no effect of IL-6 loss on the induction of myogenin, and cyclin D1 mRNA expression after 3d OV. However, MyoD mRNA expression in 3d OV IL-6-/- muscle was attenuated when compared to WT overload mice. Conclusion IL-6 appears to be necessary for the normal regulation of extracellular matrix remodeling during overload-induced growth. PMID:19681796

  17. Induction of Tenogenic Differentiation Mediated by Extracellular Tendon Matrix and Short-Term Cyclic Stretching

    PubMed Central

    Plenge, Amelie; Heller, Sandra; Pfeiffer, Bastian; Kasper, Cornelia

    2016-01-01

    Tendon and ligament pathologies are still a therapeutic challenge, due to the difficulty in restoring the complex extracellular matrix architecture and biomechanical strength. While progress is being made in cell-based therapies and tissue engineering approaches, comprehensive understanding of the fate of progenitor cells in tendon healing is still lacking. The aim of this study was to investigate the effect of decellularized tendon matrix and moderate cyclic stretching as natural stimuli which could potentially direct tenogenic fate. Equine adipose-derived mesenchymal stromal cells (MSC) were seeded on decellularized tendon matrix scaffolds. Mechanical stimulation was applied in a custom-made cyclic strain bioreactor. Assessment was performed 4 h, 8 h, and 24 h following mechanical stimulation. Scaffold culture induced cell alignment and changes in expression of tendon-related genes, although cell viability was decreased compared to monolayer culture. Short mechanical stimulation periods enhanced most of the scaffold-induced effects. Collagen 1A2 expression levels were decreased, while collagen 3A1 and decorin levels were increased. Tenascin-C and scleraxis expression showed an initial decrease but had increased 24 h after stimulation. The results obtained suggest that decellularized tendon matrix, supported by cyclic stretching, can induce tenogenic differentiation and the synthesis of tendon components important for matrix remodeling.

  18. Sca-1 expression is required for efficient remodeling of the extracellular matrix during skeletal muscle regeneration

    PubMed Central

    Kafadar, Kimberly A.; Yi, Lin; Ahmad, Yusra; So, Leslie; Rossi, Fabio; Pavlath, Grace K.

    2009-01-01

    Sca-1 (Stem Cell Antigen-1) is a member of the Ly-6 family proteins that functions in cell growth, differentiation, and self-renewal in multiple tissues. In skeletal muscle Sca-1 negatively regulates myoblast proliferation and differentiation, and may function in the maintenance of progenitor cells. We investigated the role of Sca-1 in skeletal muscle regeneration and show here that Sca-1 expression is upregulated in a subset of myogenic cells upon muscle injury. We demonstrate that extract from crushed muscle upregulates Sca-1 expression in myoblasts in vitro, and that this effect is reversible and independent of cell proliferation. Sca-1-/- mice exhibit defects in muscle regeneration, with the development of fibrosis following injury. Sca-1-/- muscle displays reduced activity of matrix metalloproteinases, critical regulators of extracellular matrix remodeling. Interestingly, we show that the number of satellite cells is similar in wild-type and Sca-1-/- muscle, suggesting that in satellite cells Sca-1 does not play a role in self-renewal. We hypothesize that Sca-1 upregulates, directly or indirectly, the activity of matrix metalloproteinases, leading to matrix breakdown and efficient muscle regeneration. Further elucidation of the role of Sca-1 in matrix remodeling may aid in the development of novel therapeutic strategies for the treatment of fibrotic diseases. PMID:19059231

  19. Local extracellular matrix alignment directs cellular protrusion dynamics and migration through Rac1 and FAK.

    PubMed

    Carey, Shawn P; Goldblatt, Zachary E; Martin, Karen E; Romero, Bethsabe; Williams, Rebecca M; Reinhart-King, Cynthia A

    2016-08-01

    Cell migration within 3D interstitial microenvironments is sensitive to extracellular matrix (ECM) properties, but the mechanisms that regulate migration guidance by 3D matrix features remain unclear. To examine the mechanisms underlying the cell migration response to aligned ECM, which is prevalent at the tumor-stroma interface, we utilized time-lapse microscopy to compare the behavior of MDA-MB-231 breast adenocarcinoma cells within randomly organized and well-aligned 3D collagen ECM. We developed a novel experimental system in which cellular morphodynamics during initial 3D cell spreading served as a reductionist model for the complex process of matrix-directed 3D cell migration. Using this approach, we found that ECM alignment induced spatial anisotropy of cells' matrix probing by promoting protrusion frequency, persistence, and lengthening along the alignment axis and suppressing protrusion dynamics orthogonal to alignment. Preference for on-axis behaviors was dependent upon FAK and Rac1 signaling and translated across length and time scales such that cells within aligned ECM exhibited accelerated elongation, front-rear polarization, and migration relative to cells in random ECM. Together, these findings indicate that adhesive and protrusive signaling allow cells to respond to coordinated physical cues in the ECM, promoting migration efficiency and cell migration guidance by 3D matrix structure. PMID:27384462

  20. Teaching the Extracellular Matrix and Introducing Online Databases within a Multidisciplinary Course with i-Cell-MATRIX: A Student-Centered Approach

    ERIC Educational Resources Information Center

    Sousa, Joao Carlos; Costa, Manuel Joao; Palha, Joana Almeida

    2010-01-01

    The biochemistry and molecular biology of the extracellular matrix (ECM) is difficult to convey to students in a classroom setting in ways that capture their interest. The understanding of the matrix's roles in physiological and pathological conditions study will presumably be hampered by insufficient knowledge of its molecular structure.…

  1. Nonlinear mechanical response of the extracellular matrix: learning from articular cartilage

    NASA Astrophysics Data System (ADS)

    Kearns, Sarah; Das, Moumita

    2015-03-01

    We study the mechanical structure-function relations in the extracellular matrix (ECM) with focus on nonlinear shear and compression response. As a model system, our study focuses on the ECM in articular cartilage tissue which has two major mechanobiological components: a network of the biopolymer collagen that acts as a stiff, reinforcing matrix, and a flexible aggrecan network that facilitates deformability. We model this system as a double network hydrogel made of interpenetrating networks of stiff and flexible biopolymers respectively. We study the linear and nonlinear mechanical response of the model ECM to shear and compression forces using a combination of rigidity percolation theory and energy minimization approaches. Our results may provide useful insights into the design principles of the ECM as well as biomimetic hydrogels that are mechanically robust and can, at the same time, easily adapt to cues in their surroundings.

  2. Embryonic lung morphogenesis in organ culture: experimental evidence for a proteoglycan function in the extracellular matrix

    NASA Technical Reports Server (NTRS)

    Spooner, B. S.; Bassett, K. E.; Spooner, B. S. Jr

    1993-01-01

    The lung rudiment, isolated from mid-gestation (11 day) mouse embryos, can undergo morphogenesis in organ culture. Observation of living rudiments, in culture, reveals both growth and ongoing bronchiolar branching activity. To detect proteoglycan (PG) biosynthesis, and deposition in the extracellular matrix, rudiments were metabolically labeled with radioactive sulfate, then fixed, embedded, sectioned and processed for autoradiography. The sulfated glycosaminoglycan (GAG) types, composing the carbohydrate component of the proteoglycans, were evaluated by selective GAG degradative approaches that showed chondroitin sulfate PG principally associated with the interstitial matrix, and heparan sulfate PG principally associated with the basement membrane. Experiments using the proteoglycan biosynthesis disrupter, beta-xyloside, suggest that when chondroitin sulfate PG deposition into the ECM is perturbed, branching morphogenesis is compromised.

  3. Quantitative analysis of 3D extracellular matrix remodelling by pancreatic stellate cells

    PubMed Central

    Robinson, Benjamin K.; Cortes, Ernesto; Rice, Alistair J.; Sarper, Muge

    2016-01-01

    ABSTRACT Extracellular matrix (ECM) remodelling is integral to numerous physiological and pathological processes in biology, such as embryogenesis, wound healing, fibrosis and cancer. Until recently, most cellular studies have been conducted on 2D environments where mechanical cues significantly differ from physiologically relevant 3D environments, impacting cellular behaviour and masking the interpretation of cellular function in health and disease. We present an integrated methodology where cell-ECM interactions can be investigated in 3D environments via ECM remodelling. Monitoring and quantification of collagen-I structure in remodelled matrices, through designated algorithms, show that 3D matrices can be used to correlate remodelling with increased ECM stiffness observed in fibrosis. Pancreatic stellate cells (PSCs) are the key effectors of the stromal fibrosis associated to pancreatic cancer. We use PSCs to implement our methodology and demonstrate that PSC matrix remodelling capabilities depend on their contractile machinery and β1 integrin-mediated cell-ECM attachment. PMID:27170254

  4. Synthetic Mimics of the Extracellular Matrix: How Simple is Complex Enough?

    PubMed Central

    Kyburz, Kyle A.; Anseth, Kristi S.

    2015-01-01

    Cells reside in a complex and dynamic extracellular matrix where they interact with a myriad of biophysical and biochemical cues that direct their function and regulate tissue homeostasis, wound repair, and even pathophysiological events. There is a desire in the biomaterials community to develop synthetic hydrogels to recapitulate facets of the ECM for in vitro culture platforms and tissue engineering applications. Advances in synthetic hydrogel design and chemistries, including user-tunable platforms, have broadened the field’s understanding of the role of matrix cues in directing cellular processes and enabled the design of improved tissue engineering scaffolds. This review focuses on recent advances in the development and fabrication of hydrogels and discusses what aspects of ECM signals can be incorporated to direct cell function in different contexts. PMID:25753017

  5. The planar cell polarity protein VANGL2 coordinates remodeling of the extracellular matrix.

    PubMed

    Williams, B Blairanne; Mundell, Nathan; Dunlap, Julie; Jessen, Jason

    2012-07-01

    Understanding how planar cell polarity (PCP) is established, maintained, and coordinated in migrating cell populations is an important area of research with implications for both embryonic morphogenesis and tumor cell invasion. We recently reported that the PCP protein Vang-like 2 (VANGL2) regulates the endocytosis and cell surface level of membrane type-1 matrix metalloproteinase (MMP14 or MT1-MMP). Here, we further discuss these findings in terms of extracellular matrix (ECM) remodeling, cell migration, and zebrafish gastrulation. We also demonstrate that VANGL2 function impacts the focal degradation of ECM by human cancer cells including the formation or stability of invadopodia. Together, our findings implicate MMP14 as a downstream effector of VANGL2 signaling and suggest a model whereby the regulation of pericellular proteolysis is a fundamental aspect of PCP in migrating cells. PMID:23060953

  6. The planar cell polarity protein VANGL2 coordinates remodeling of the extracellular matrix

    PubMed Central

    Williams, B. Blairanne; Mundell, Nathan; Dunlap, Julie; Jessen, Jason

    2012-01-01

    Understanding how planar cell polarity (PCP) is established, maintained, and coordinated in migrating cell populations is an important area of research with implications for both embryonic morphogenesis and tumor cell invasion. We recently reported that the PCP protein Vang-like 2 (VANGL2) regulates the endocytosis and cell surface level of membrane type-1 matrix metalloproteinase (MMP14 or MT1-MMP). Here, we further discuss these findings in terms of extracellular matrix (ECM) remodeling, cell migration, and zebrafish gastrulation. We also demonstrate that VANGL2 function impacts the focal degradation of ECM by human cancer cells including the formation or stability of invadopodia. Together, our findings implicate MMP14 as a downstream effector of VANGL2 signaling and suggest a model whereby the regulation of pericellular proteolysis is a fundamental aspect of PCP in migrating cells. PMID:23060953

  7. Thrombopoietin/TGF-β1 Loop Regulates Megakaryocyte Extracellular Matrix Component Synthesis.

    PubMed

    Abbonante, Vittorio; Di Buduo, Christian A; Gruppi, Cristian; Malara, Alessandro; Gianelli, Umberto; Celesti, Giuseppe; Anselmo, Achille; Laghi, Luigi; Vercellino, Marco; Visai, Livia; Iurlo, Alessandra; Moratti, Remigio; Barosi, Giovanni; Rosti, Vittorio; Balduini, Alessandra

    2016-04-01

    Extracellular matrix (ECM) components initiate crucial biochemical and biomechanical cues that are required for bone marrow homeostasis. In our research, we prove that a peri-cellular matrix composed primarily of type III and type IV collagens, and fibronectin surrounds human megakaryocytes in the bone marrow. The data we collected support the hypothesis that bone marrow megakaryocytes possess a complete mechanism to synthesize the ECM components, and that thrombopoietin is a pivotal regulator of this new function inducing transforming growth factor-β1 (TGF-β1) release and consequent activation of the downstream pathways, both in vitro and in vivo. This activation results in a dose dependent increase of ECM component synthesis by megakaryocytes, which is reverted upon incubation with JAK and TGF-β1 receptor specific inhibitors. These data are pivotal for understanding the central role of megakaryocytes in creating their own regulatory niche within the bone marrow environment. Stem Cells 2016;34:1123-1133. PMID:26748484

  8. Development of biomimetic nanocomposites as bone extracellular matrix for human osteoblastic cells.

    PubMed

    Bhowmick, Arundhati; Mitra, Tapas; Gnanamani, Arumugam; Das, Manas; Kundu, Patit Paban

    2016-05-01

    Here, we have developed biomimetic nanocomposites containing chitosan, poly(vinyl alcohol) and nano-hydroxyapatite-zinc oxide as bone extracellular matrix for human osteoblastic cells and characterized by Fourier transform infrared spectroscopy, powder X-ray diffraction. Scanning electron microscopy images revealed interconnected macroporous structures. Moreover, in this study, the problem related to fabricating a porous composite with good mechanical strength has been resolved by incorporating 5wt% of nano-hydroxyapatite-zinc oxide into chitosan-poly(vinyl alcohol) matrix; the present composite showed high tensile strength (20.25MPa) while maintaining appreciable porosity (65.25%). These values are similar to human cancellous bone. These nanocomposites also showed superior water uptake, antimicrobial and biodegradable properties than the previously reported results. Compatibility with human blood and pH was observed, indicating nontoxicity of these materials to the human body. Moreover, proliferation of osteoblastic MG-63 cells onto the nanocomposites was also observed without having any negative effect. PMID:26876999

  9. Dilation and degradation of the brain extracellular matrix enhances penetration of infused polymer nanoparticles

    PubMed Central

    Neeves, Keith B.; Sawyer, Andrew J.; Foley, Conor P.; Saltzman, W. Mark; Olbricht, William L.

    2007-01-01

    This study investigates methods of manipulating the brain extracellular matrix (ECM) to enhance the penetration of nanoparticle drug carriers in convection-enhanced delivery (CED). A probe was fabricated with two independent microfluidic channels to infuse, either simultaneously or sequentially, nanoparticles and ECM-modifying agents. Infusions were performed in the striatum of the normal rat brain. Monodisperse polystyrene particles with a diameter of 54 nm were used as a model nanoparticle system. Because the size of these particles is comparable to the effective pore size of the ECM, their transport may be significantly hindered compared with the transport of low molecular weight molecules. To enhance the transport of the infused nanoparticles, we attempted to increase the effective pore size of the ECM by two methods: dilating the extracellular space and degrading selected constituents of the ECM. Two methods of dilating the extracellular space were investigated: co-infusion of nanoparticles and a hyperosmolar solution of mannitol, and pre-infusion of an isotonic buffer solution followed by infusion of nanoparticles. These treatments resulted in an increase in the nanoparticle distribution volume of 50% and 123%, respectively. To degrade hyaluronan, a primary structural component of the brain ECM, a pre-infusion of hyaluronidase (20,000 U/mL) was followed after 30 min by infusion of nanoparticles. This treatment resulted in an increase in the nanoparticle distribution of 64%. Our results suggest that both dilation and enzymatic digestion can be incorporated into CED protocols to enhance nanoparticle penetration. PMID:17920047

  10. Aldosterone and myocardial extracellular matrix expansion in type 2 diabetes mellitus.

    PubMed

    Rao, Ajay D; Shah, Ravi V; Garg, Rajesh; Abbasi, Siddique A; Neilan, Tomas G; Perlstein, Todd S; Di Carli, Marcelo F; Jerosch-Herold, Michael; Kwong, Raymond Y; Adler, Gail K

    2013-07-01

    Myocardial extracellular matrix expansion and reduced coronary flow reserve (CFR) occur in patients with type 2 diabetes mellitus without heart failure or coronary artery disease. Because aldosterone is implicated in the pathophysiology of cardiac fibrosis and vascular injury, the aim of this study was to test the hypothesis that aldosterone is associated with extracellular matrix expansion and reduced CFR in type 2 diabetes mellitus. Patients with type 2 diabetes mellitus without evidence of coronary artery disease were recruited. Blood pressure, lipid management, and glycemic control were optimized over 3 months. Cardiac magnetic resonance imaging with T1 mapping was used to measure myocardial extracellular volume (ECV). Cardiac positron emission tomography was used to assess CFR. On a liberal, 250 mEq/day sodium diet, 24-hour urinary aldosterone and change in serum aldosterone with angiotensin II stimulation were measured. Fifty-three participants with type 2 diabetes (68% men, mean age 53 ± 7 years, mean body mass index 32.2 ± 4.3 kg/m², mean glycosylated hemoglobin 6.8 ± 0.7%, mean systolic blood pressure 126 ± 14 mm Hg) without infarction or ischemia by cardiac magnetic resonance and positron emission tomography were studied. Subjects had impaired CFR (2.51 ± 0.83) and elevated ECV (0.36 ± 0.05), despite normal echocardiographic diastolic function and normal left ventricular function. Myocardial ECV, but not CFR, was positively associated with 24-hour urinary aldosterone excretion (r = 0.37, p = 0.01) and angiotensin II-stimulated aldosterone increase (r = 0.35, p = 0.02). In a best-overall multivariate model (including age, gender, body mass index, glycosylated hemoglobin, and blood pressure), 24-hour urinary aldosterone was the strongest predictor of myocardial ECV (p = 0.004). In conclusion, in patients with type 2 diabetes mellitus without coronary artery disease, aldosterone is associated with myocardial extracellular matrix expansion. These

  11. Amyloid β-Protein as a Substrate Interacts with Extracellular Matrix to Promote Neurite Outgrowth

    NASA Astrophysics Data System (ADS)

    Koo, Edward H.; Park, Lisa; Selkoe, Dennis J.

    1993-05-01

    Progressive deposition of amyloid β-protein (Aβ) in brain parenchyma and blood vessels is a characteristic feature of Alzheimer disease. Recent evidence suggests that addition of solubilized synthetic Aβ to medium may produce toxic or trophic effects on cultured hippocampal neurons. Because soluble Aβ may not accumulate in significant quantities in brain, we asked whether immobilized Aβ peptide as a substrate alters neurite outgrowth from cultured rat peripheral sensory neurons. This paradigm may closely mimic the conditions in Alzheimer disease brain tissue, in which neurites contact insoluble, extracellular aggregates of β-amyloid. We detected no detrimental effects of Aβ substrate on neurite outgrowth. Rather, Aβ in combination with low doses of laminin or fibronectin enhanced neurite out-growth from these neuronal explants. Our results suggest that insoluble Aβ in the cerebral neuropil may serve as a neurite-promoting matrix, perhaps explaining the apparent regenerative response of neurites observed around amyloid plaques in Alzheimer disease. Moreover, in concert with the recent discovery of Aβ production by cultured neurons, our data suggest that Aβ plays a normal physiological role in brain by complexing with the extracellular matrix.

  12. Aortic Wall Extracellular Matrix Proteins Correlate with Syntax Score in Patients Undergoing Coronary Artery Bypass Surgery

    PubMed Central

    Chiong, Terri; Cheow, Esther S. H.; Woo, Chin C.; Lin, Xiao Y.; Khin, Lay W.; Lee, Chuen N.; Hartman, Mikael; Sze, Siu K.; Sorokin, Vitaly A.

    2016-01-01

    Aims: The SYNTAX score correlate with major cardiovascular events post-revascularization, although the histopathological basis is unclear. We aim to evaluate the association between syntax score and extracellular matrix histological characteristics of aortic punch tissue obtained during coronary artery bypass surgery (CABG). This analysis compares coronary artery bypass surgery patients with High and Low syntax score which were followed up for one year period. Methods and Results: Patients with High (score ≥ 33, (n=77)) and Low Syntax Scores (score ≤ 22, (n=71)) undergoing elective CABG were recruited prospectively. Baseline clinical characteristics and surgical risks were well matched. At 1 year, EMACCE (Sum of cardiovascular death, stroke, congestive cardiac failure, and limb, gut and myocardial ischemia) was significantly elevated in the High syntax group (P=0.022). Mass spectrometry (MS)-based quantitative iTRAQ proteomic results validated on independent cohort by immunohistochemistry (IHC) revealed that the High syntax group had significantly upraised Collagen I (P<0.0001) and Elastin (P<0.0001) content in ascending aortic wall. Conclusion: This study shows that aortic extracellular matrix (ECM) differ between High and Low syntax groups with up-regulation of Collagen I and Elastin level in High Syntax Score group. This identifies aortic punches collected during CABG as another biomarker source related with atherosclerosis severity and possible clinical outcome. PMID:27347220

  13. Brevican-containing perineuronal nets of extracellular matrix in dissociated hippocampal primary cultures.

    PubMed

    John, Nora; Krügel, Hans; Frischknecht, Renato; Smalla, Karl-Heinz; Schultz, Christian; Kreutz, Michael R; Gundelfinger, Eckart D; Seidenbecher, Constanze I

    2006-04-01

    Perineuronal nets (PNN) are specialized extracellular matrix structures enwrapping CNS neurons, which are important regulators for neuronal and synaptic functions. Brevican, a chondroitin sulfate proteoglycan, is an integral component of PNN. Here, we have investigated the appearance of these structures in hippocampal primary cultures. The expression profile of brevican in mixed cultures resembles the in vivo pattern with a strong upregulation of all isoforms during the second and 3rd weeks in culture. Brevican is primarily synthesized by co-cultured glial fibrillary acidic protein (GFAP-)-positive astrocytes and co-assembles with its interaction partners in PNN-like structures on neuronal somata and neurites as identified by counterstaining with the PNN marker Vicia villosa lectin. Both excitatory and inhibitory synapses are embedded into PNN. Furthermore, axon initial segments are strongly covered by a dense brevican coat. Altogether, we show that mature primary cultures can form PNN, and that basic features of these extracellular matrix structures may be studied in vitro. PMID:16503162

  14. Presence of extracellular DNA in the Candida albicans biofilm matrix and its contribution to biofilms

    PubMed Central

    Martins, Margarida; Uppuluri, Priya; Thomas, Derek P.; Cleary, Ian A.; Henriques, Mariana; Lopez-Ribot, José L.; Oliveira, Rosário

    2014-01-01

    DNA has been described as a structural component of the extracellular matrix (ECM) in bacterial biofilms. In Candida albicans there is a scarce knowledge concerning the contribution of extracellular DNA (eDNA) to biofilm matrix and overall structure. This work examined the presence and quantified the amount of eDNA in C.albicans biofilm ECM and the effect of DNase treatment and the addition of exogenous DNA on C. albicans biofilm development as indicators of a role for eDNA in biofilm development. We were able to detect the accumulation of eDNA in biofilm ECM extracted from C. albicans biofilms formed under conditions of flow, although the quantity of eDNA detected differed according to growth conditions, in particular with regards to the medium used to grow the biofilms. Experiments with C. albicans biofilms formed statically using a microtiter plate model indicated that the addition of exogenous DNA (>160 ng/ml) increases biofilm biomass and, conversely, DNase treatment (>0.03 mg/ml) decreases biofilm biomass at later time points of biofilm development. We present evidence for the role of eDNA in C. albicans biofilm structure and formation, consistent with eDNA being a key element of the ECM in mature C. albicans biofilms and playing a predominant role in biofilm structural integrity and maintenance. PMID:20012895

  15. Core-shell hydrogel beads with extracellular matrix for tumor spheroid formation.

    PubMed

    Yu, L; Grist, S M; Nasseri, S S; Cheng, E; Hwang, Y-C E; Ni, C; Cheung, K C

    2015-03-01

    Creating multicellular tumor spheroids is critical for characterizing anticancer treatments since they may provide a better model of the tumor than conventional monolayer culture. Moreover, tumor cell interaction with the extracellular matrix can determine cell organization and behavior. In this work, a microfluidic system was used to form cell-laden core-shell beads which incorporate elements of the extracellular matrix and support the formation of multicellular spheroids. The bead core (comprising a mixture of alginate, collagen, and reconstituted basement membrane, with gelation by temperature control) and shell (comprising alginate hydrogel, with gelation by ionic crosslinking) were simultaneously formed through flow focusing using a cooled flow path into the microfluidic chip. During droplet gelation, the alginate acts as a fast-gelling shell which aids in preventing droplet coalescence and in maintaining spherical droplet geometry during the slower gelation of the collagen and reconstituted basement membrane components as the beads warm up. After droplet gelation, the encapsulated MCF-7 cells proliferated to form uniform spheroids when the beads contained all three components: alginate, collagen, and reconstituted basement membrane. The dose-dependent response of the MCF-7 cell tumor spheroids to two anticancer drugs, docetaxel and tamoxifen, was compared to conventional monolayer culture. PMID:25945144

  16. Bubaline Cholecyst Derived Extracellular Matrix for Reconstruction of Full Thickness Skin Wounds in Rats

    PubMed Central

    Shakya, Poonam; Sharma, A. K.; Kumar, Naveen; Vellachi, Remya; Mathew, Dayamon D.; Dubey, Prasoon; Singh, Kiranjeet; Shrivastava, Sonal; Shrivastava, Sameer; Maiti, S. K.; Hasan, Anwarul; Singh, K. P.

    2016-01-01

    An acellular cholecyst derived extracellular matrix (b-CEM) of bubaline origin was prepared using anionic biological detergent. Healing potential of b-CEM was compared with commercially available collagen sheet (b-CS) and open wound (C) in full thickness skin wounds in rats. Thirty-six clinically healthy adult Sprague Dawley rats of either sex were randomly divided into three equal groups. Under general anesthesia, a full thickness skin wound (20 × 20 mm2) was created on the dorsum of each rat. The defect in group I was kept as open wound and was taken as control. In group II, the defect was repaired with commercially available collagen sheet (b-CS). In group III, the defect was repaired with cholecyst derived extracellular matrix of bovine origin (b-CEM). Planimetry, wound contracture, and immunological and histological observations were carried out to evaluate healing process. Significantly (P < 0.05) increased wound contraction was observed in b-CEM (III) as compared to control (I) and b-CS (II) on day 21. Histologically, improved epithelization, neovascularization, fibroplasia, and best arranged collagen fibers were observed in b-CEM (III) as early as on postimplantation day 21. These findings indicate that b-CEM have potential for biomedical applications for full thickness skin wound repair in rats. PMID:27127678

  17. Trafficking Mechanisms of Extracellular Matrix Macromolecules: Insights from Vertebrate Development and Human Diseases

    PubMed Central

    Unlu, Gokhan; Levic, Daniel S.; Melville, David B.; Knapik, Ela W.

    2014-01-01

    Cellular life depends on protein transport and membrane traffic. In multicellular organisms, membrane traffic is required for extracellular matrix deposition, cell adhesion, growth factor release, and receptor signaling, which are collectively required to integrate the development and physiology of tissues and organs. Understanding the regulatory mechanisms that govern cargo and membrane flow presents a prime challenge in cell biology. Extracellular matrix (ECM) secretion remains poorly understood, although given its essential roles in the regulation of cell migration, differentiation, and survival, ECM secretion mechanisms are likely to be tightly controlled. Recent studies in vertebrate model systems, from fishes to mammals and in human patients, have revealed complex and diverse loss-of-function phenotypes associated with mutations in components of the secretory machinery. A broad spectrum of diseases from skeletal and cardiovascular to neurological deficits have been linked to ECM trafficking. These discoveries have directly challenged the prevailing view of secretion as an essential but monolithic process. Here, we will discuss the latest findings on mechanisms of ECM trafficking in vertebrates. PMID:24333299

  18. Salvage of failed local and regional flaps with porcine urinary bladder extracellular matrix aided tissue regeneration.

    PubMed

    Kruper, Gregory J; Vandegriend, Zachary P; Lin, Ho-Sheng; Zuliani, Giancarlo F

    2013-01-01

    Local and regional flap failure can be a major complication in head and neck surgery, which continue to be prevalent for a number of reasons including poor flap design, improper surgical technique, and poor tissue vascularity. Dealing with these failures can be quite difficult. Surgical debridement, flap revisions, and complex wound regimens are necessitated to reestablish appropriate tissue coverage. Traditional use of wet to dry dressing to enable proper wound granulation and possible closure with additional flaps or skin grafts is a laborious process. Such treatments place great time burdens on the patient, physicians, and nurses. Because the face and neck possess a complex three-dimensional topography, wound dressings are inherently complex to design and change. Many patients also require postoperative treatments such as radiation and chemotherapy to treat aggressive malignancies, and delay in wound healing leads to a delay in adjuvant treatment. Recently, advances in regenerative medicine, specifically xenogeneic extracellular matrix compounds, have been shown to promote tissue growth while limiting scar tissue formation (Badylak 2004). To our knowledge, this paper is the first case series using the porcine extracellular matrix bioscaffold (MatriStem ACell, Columbia, MD, USA) to salvage flaps with extensive wound breakdown on the face and neck. PMID:24191216

  19. Up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver

    PubMed Central

    Zhang, Shuai; Li, Tao-Sheng; Soyama, Akihiko; Tanaka, Takayuki; Yan, Chen; Sakai, Yusuke; Hidaka, Masaaki; Kinoshita, Ayaka; Natsuda, Koji; Fujii, Mio; Kugiyama, Tota; Baimakhanov, Zhassulan; Kuroki, Tamotsu; Gu, Weili; Eguchi, Susumu

    2016-01-01

    Although the healthy liver is known to have high regenerative potential, poor liver regeneration under pathological conditions remains a substantial problem. We investigated the key molecules that impair the regeneration of cholestatic liver. C57BL/6 mice were randomly subjected to partial hepatectomy and bile duct ligation (PH+BDL group, n = 16), partial hepatectomy only (PH group, n = 16), or sham operation (Sham group, n = 16). The liver sizes and histological findings were similar in the PH and sham groups 14 days after operation. However, compared with those in the sham group, the livers in mice in the PH+BDL group had a smaller size, a lower cell proliferative activity, and more fibrotic tissue 14 days after the operation, suggesting the insufficient regeneration of the cholestatic liver. Pathway-focused array analysis showed that many genes were up- or down-regulated over 1.5-fold in both PH+BDL and PH groups at 1, 3, 7, and 14 days after treatment. Interestingly, more genes that were functionally related to the extracellular matrix and inflammatory chemokines were found in the PH+BDL group than in the PH group at 7 and 14 days after treatment. Our data suggest that up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver. PMID:27226149

  20. Up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver.

    PubMed

    Zhang, Shuai; Li, Tao-Sheng; Soyama, Akihiko; Tanaka, Takayuki; Yan, Chen; Sakai, Yusuke; Hidaka, Masaaki; Kinoshita, Ayaka; Natsuda, Koji; Fujii, Mio; Kugiyama, Tota; Baimakhanov, Zhassulan; Kuroki, Tamotsu; Gu, Weili; Eguchi, Susumu

    2016-01-01

    Although the healthy liver is known to have high regenerative potential, poor liver regeneration under pathological conditions remains a substantial problem. We investigated the key molecules that impair the regeneration of cholestatic liver. C57BL/6 mice were randomly subjected to partial hepatectomy and bile duct ligation (PH+BDL group, n = 16), partial hepatectomy only (PH group, n = 16), or sham operation (Sham group, n = 16). The liver sizes and histological findings were similar in the PH and sham groups 14 days after operation. However, compared with those in the sham group, the livers in mice in the PH+BDL group had a smaller size, a lower cell proliferative activity, and more fibrotic tissue 14 days after the operation, suggesting the insufficient regeneration of the cholestatic liver. Pathway-focused array analysis showed that many genes were up- or down-regulated over 1.5-fold in both PH+BDL and PH groups at 1, 3, 7, and 14 days after treatment. Interestingly, more genes that were functionally related to the extracellular matrix and inflammatory chemokines were found in the PH+BDL group than in the PH group at 7 and 14 days after treatment. Our data suggest that up-regulated extracellular matrix components and inflammatory chemokines may impair the regeneration of cholestatic liver. PMID:27226149

  1. Core-shell hydrogel beads with extracellular matrix for tumor spheroid formation

    PubMed Central

    Yu, L.; Grist, S. M.; Nasseri, S. S.; Ni, C.; Cheung, K. C.

    2015-01-01

    Creating multicellular tumor spheroids is critical for characterizing anticancer treatments since they may provide a better model of the tumor than conventional monolayer culture. Moreover, tumor cell interaction with the extracellular matrix can determine cell organization and behavior. In this work, a microfluidic system was used to form cell-laden core-shell beads which incorporate elements of the extracellular matrix and support the formation of multicellular spheroids. The bead core (comprising a mixture of alginate, collagen, and reconstituted basement membrane, with gelation by temperature control) and shell (comprising alginate hydrogel, with gelation by ionic crosslinking) were simultaneously formed through flow focusing using a cooled flow path into the microfluidic chip. During droplet gelation, the alginate acts as a fast-gelling shell which aids in preventing droplet coalescence and in maintaining spherical droplet geometry during the slower gelation of the collagen and reconstituted basement membrane components as the beads warm up. After droplet gelation, the encapsulated MCF-7 cells proliferated to form uniform spheroids when the beads contained all three components: alginate, collagen, and reconstituted basement membrane. The dose-dependent response of the MCF-7 cell tumor spheroids to two anticancer drugs, docetaxel and tamoxifen, was compared to conventional monolayer culture. PMID:25945144

  2. The interaction of Tamm-Horsfall protein with the extracellular matrix.

    PubMed Central

    Lambert, C; Brealey, R; Steele, J; Rook, G A

    1993-01-01

    Tamm-Horsfall protein (THP) is the major glycoprotein component of urine, yet its biological role remains obscure. Recent reports have suggested that a concanavalin A (Con A)-binding fraction of THP from pregnancy urine can bind the cytokines tumour necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL-1). In order to investigate this claim in relation to THP from normal adult urine we raised monoclonal antibodies to THP and sought THP/TNF-alpha interactions in three separate assay systems. We found no evidence that THP binds to TNF-alpha under physiological conditions, but we observed that it exerts a weak, probably not physiologically relevant, but reproducible inhibitory effect on the toxicity of TNF-alpha for monolayers of L929 cells, even when the cells are pretreated with the THP, and washed before addition of the cytokine. Since our preparations of THP do not interact directly with TNF-alpha we postulated an interaction with the cells themselves, or with their extracellular matrix. The THP was found by ELISA, immunoblotting and immunohistology, to bind to as yet unidentified components of the extracellular matrix in a manner dependent on cations, pH and carbohydrates. These data, considered in the light of the published amino acid sequence and biochemical properties, suggest that THP is a member of a structural glycoprotein family known to modulate cell adhesion. Images Figure 4 Figure 5 PMID:8344699

  3. Salvage of Failed Local and Regional Flaps with Porcine Urinary Bladder Extracellular Matrix Aided Tissue Regeneration

    PubMed Central

    Kruper, Gregory J.; VandeGriend, Zachary P.; Lin, Ho-Sheng; Zuliani, Giancarlo F.

    2013-01-01

    Local and regional flap failure can be a major complication in head and neck surgery, which continue to be prevalent for a number of reasons including poor flap design, improper surgical technique, and poor tissue vascularity. Dealing with these failures can be quite difficult. Surgical debridement, flap revisions, and complex wound regimens are necessitated to reestablish appropriate tissue coverage. Traditional use of wet to dry dressing to enable proper wound granulation and possible closure with additional flaps or skin grafts is a laborious process. Such treatments place great time burdens on the patient, physicians, and nurses. Because the face and neck possess a complex three-dimensional topography, wound dressings are inherently complex to design and change. Many patients also require postoperative treatments such as radiation and chemotherapy to treat aggressive malignancies, and delay in wound healing leads to a delay in adjuvant treatment. Recently, advances in regenerative medicine, specifically xenogeneic extracellular matrix compounds, have been shown to promote tissue growth while limiting scar tissue formation (Badylak 2004). To our knowledge, this paper is the first case series using the porcine extracellular matrix bioscaffold (MatriStem ACell, Columbia, MD, USA) to salvage flaps with extensive wound breakdown on the face and neck. PMID:24191216

  4. Response of Inflammatory Mediators, Extracellular Matrix Proteins and Stem and Progenitor Cells to Emphysema.

    PubMed

    Skurikhin, E G; Pakhomova, A V; Krupin, V A; Pershina, O V; Pan, E S; Ermolaeva, L A; Vaizova, O E; Rybalkina, O Yu; Dygai, A M

    2016-08-01

    Inflammation, extracellular matrix proteins (hydroxyproline, connective tissue growth factor, collagen, and fibronectin), stem and progenitor cells (multipotent mesenchymal stromal cells, Clara cells, angiogenesis, precursors, endothelial and epithelial cells) were studied in female C57Bl/6 mice with experimental elastase-induced emphysema. Diffuse emphysema reduced the number of endothelial (CD45(-)CD31(+)CD34(+)) and epithelial (CD45(-)CD117(+)CD49f(+)) cells, induced microcirculation disturbances, and decreased the area occupied by the connective tissue. Emphysematous changes in the lungs were accompanied by infiltration of the alveolar septa with macrophages and lymphocytes, increase in the serum and lung concentrations of transforming growth factor-β, IL-1β, IL-2, IL-5, IL-10, and IL-13, and lung concentration of IL-17. In the lungs, inflammation was associated with marked increase in the number of multipotent mesenchymal stromal cells CD90(+)CD73(+)CD106(+)CD44(+)) and Clara cells (CD45(-)CD34(-)CD31(-)Sca1(+)) and overexpression of extracellular matrix proteins (hydroxyproline, connective tissue growth factor, collagen, fibronectin) and Clara cells protein. On the other hand, elastase reduced the number of angiogenic precursor cells (CD45(-)CD117(+)Flk1(+)). PMID:27591877

  5. Interleukin 18 and extracellular matrix metalloproteinase inducer cross-regulation: implications in acute myocardial infarction.

    PubMed

    Xie, Shuang-Lun; Chen, Yu-Yang; Zhang, Hai-Feng; Deng, Bing-Qing; Shu, Xiao-Rong; Su, Zi-Zhuo; Lin, Yong-Qing; Nie, Ru-Qiong; Wang, Jing-Feng

    2015-03-01

    Circulating interleukin-18 (IL-18) is thought to promote atherosclerosis and cardiovascular complications such as plaque rupture. Atherosclerosis is also characterized by smooth muscle cell migration, a consequence of extracellular matrix (ECM) degradation regulated by metalloproteinases (MMPs). Because extracellular matrix metalloproteinase inducer (EMMPRIN) has been shown to promote plaque instability by inducing ECM degradation and MMP synthesis, we investigated whether a cross-regulatory interaction exists between IL-18 and EMMPRIN in human monocytes. EMMPRIN levels in monocytes were markedly greater in 20 patients with acute myocardial infarction (AMI) compared with 20 patients with stable angina pectoris or 20 healthy volunteers (control group). The levels of IL-18 and MMP-9 in serum were also significantly greater in the AMI group in comparison with the other 2 groups. IL-18 levels positively correlated with increased levels of EMMPRIN in monocytes. In vitro, the expression of EMMPRIN was increased in monocytes cultured with IL-18, and IL-18 secretion was augmented in monocytes cultured with EMMPRIN. Gene silencing of EMMPRIN by small interfering RNA reduced monocyte secretion of both IL-18 and MMP-9. In the present study, cross-regulation between IL-18 and EMMPRIN in monocytes was demonstrated. This interaction may amplify the inflammatory cascade and be responsible for increased monocytic MMP-9 serum levels in atherosclerosis, contributing to atherosclerotic plaque destabilization and subsequent AMI. PMID:25267095

  6. Promoting external inosculation of prevascularised tissue constructs by pre-cultivation in an angiogenic extracellular matrix.

    PubMed

    Laschke, M W; Mussawy, H; Schuler, S; Eglin, D; Alini, M; Menger, M D

    2010-01-01

    The engineering of preformed microvessels offers the promising opportunity to rapidly vascularise implanted tissue constructs by the process of inosculation. Herein, we analyzed whether this process may further be accelerated by cultivation of prevascularised tissue constructs in Matrigel before implantation. Nano-size hydroxyapatite particles/poly(ester-urethane) scaffolds were implanted into the flank of FVB/N-TgN (Tie2/GFP) 287 Sato mice to allow the ingrowth of a granulation tissue with green fluorescent protein (GFP)-positive blood vessels. After harvesting, these prevascularised constructs were then transferred into dorsal skinfold chambers of FVB/N recipient mice to study the process of inosculation. The constructs were implanted directly after embedding in Matrigel or after 3 days of cultivation in the extracellular matrix. Matrigel-free constructs served as control. Cultivation in Matrigel resulted in the outgrowth of CD31/GFP-positive vascular sprouts. Vascularisation of these constructs was markedly improved when compared to the other two groups, as indicated by a significantly elevated functional microvessel density between days 6 to 14 after implantation into the dorsal skinfold chamber. This was associated with an increased number of GFP-positive blood vessels growing into the surrounding host tissue. Thus, the blood supply to prevascularised tissue constructs can be accelerated by their pre-cultivation in an angiogenic extracellular matrix, promoting external inosculation of the preformed microvascular networks with the host microvasculature. PMID:21154242

  7. In the presence of danger: the extracellular matrix defensive response to central nervous system injury

    PubMed Central

    Jakeman, Lyn B.; Williams, Kent E.; Brautigam, Bryan

    2014-01-01

    Glial cells in the central nervous system (CNS) contribute to formation of the extracellular matrix, which provides adhesive sites, signaling molecules, and a diffusion barrier to enhance efficient neurotransmission and axon potential propagation. In the normal adult CNS, the extracellular matrix (ECM) is relatively stable except in selected regions characterized by dynamic remodeling. However, after trauma such as a spinal cord injury or cortical contusion, the lesion epicenter becomes a focus of acute neuroinflammation. The activation of the surrounding glial cells leads to a dramatic change in the composition of the ECM at the edges of the lesion, creating a perilesion environment dominated by growth inhibitory molecules and restoration of the peripheral/central nervous system border. An advantage of this response is to limit the invasion of damaging cells and diffusion of toxic molecules into the spared tissue regions, but this occurs at the cost of inhibiting migration of endogenous repair cells and preventing axonal regrowth. The following review was prepared by reading and discussing over 200 research articles in the field published in PubMed and selecting those with significant impact and/or controversial points. This article highlights structural and functional features of the normal adult CNS ECM and then focuses on the reactions of glial cells and changes in the perilesion border that occur following spinal cord or contusive brain injury. Current research strategies directed at modifying the inhibitory perilesion microenvironment without eliminating the protective functions of glial cell activation are discussed. PMID:24999352

  8. Solubilized liver extracellular matrix maintains primary rat hepatocyte phenotype in-vitro.

    PubMed

    Loneker, Abigail E; Faulk, Denver M; Hussey, George S; D'Amore, Antonio; Badylak, Stephen F

    2016-04-01

    Whole organ engineering and cell-based regenerative medicine approaches are being investigated as potential therapeutic options for end-stage liver failure. However, a major challenge of these strategies is the loss of hepatic specific function after hepatocytes are removed from their native microenvironment. The objective of the present study was to determine if solubilized liver extracellular matrix (ECM), when used as a media supplement, can better maintain hepatocyte phenotype compared to type I collagen alone or solubilized ECM harvested from a non-liver tissue source. Liver extracellular matrix (LECM) from four different species was isolated via liver tissue decellularization, solubilized, and then used as a media supplement for primary rat hepatocytes (PRH). The four species of LECM investigated were human, porcine, canine and rat. Cell morphology, albumin secretion, and ammonia metabolism were used to assess maintenance of hepatocyte phenotype. Biochemical and mechanical characterization of each LECM were also conducted. Results showed that PRH's supplemented with canine and porcine LECM maintained their phenotype to a greater extent compared to all other groups. PRH's supplemented with canine and porcine LECM showed increased bile production, increased albumin production, and the formation of multinucleate cells. The findings of the present study suggest that solubilized liver ECM can support in-vitro hepatocyte culture and should be considered for therapeutic and diagnostic techniques that utilize hepatocytes. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 957-965, 2016. PMID:26704367

  9. Science of Hyaluronic Acid Beyond Filling: Fibroblasts and Their Response to the Extracellular Matrix.

    PubMed

    Landau, Marina; Fagien, Steven

    2015-11-01

    Loss of viscoelasticity is one of the primarily signs of skin aging, followed by appearance of visible wrinkles. Hyaluronic acid (HA)-based fillers are widely used to fill wrinkles and compensate for volume loss. Recent clinical observations demonstrate persistence of the filling effect longer than the biological availability of the filler. Stimulation of new collagen by cross-linked HA and up-regulation of elastin have been suggested as possible explanation to this observation and have been supported experimentally. Cross-linked HA substitutes for fragmented collagen in restoring extracellular matrix required for normal activity of fibroblasts, such as collagen and elastin production. To restore extracellular matrix efficiently, serial monthly treatments are required. Boosting of facial and nonfacial skin through fibroblast activation is a new indication for HA-based products. Injectable HA has also been recently registered in Europe as agents specific for the improvement of skin quality (Restylane Skinboosters). Further explanation of the possible mechanisms supported by long-term clinical examples is presented herein. PMID:26441098

  10. Expression of extracellular matrix metalloproteinase inducer (EMMPRIN) and its related extracellular matrix degrading enzymes in the endometrium during estrous cycle and early gestation in cattle

    PubMed Central

    2010-01-01

    Background Extracellular matrix metalloproteinase inducer (EMMPRIN) regulates several biological functions involving the modulation of cell behaviors via cell-cell and cell-matrix interactions. According to its diverse functions, we hypothesized that EMMPRIN may play an important role in endometrial remodeling and establishment of pregnancy in cow. Methods In this study, endometrial tissues from the cyclic cows during before ovulation, after ovulation and middle of estrous cycle; and pregnant endometrial tissues from Day 19 to 35 of gestation have been used. Expression of mRNA was analyzed by RT-PCR, qPCR and in situ hybridization whereas protein expression by immunohistochemistry and western blot analysis. Results EMMPRIN mRNA was expressed in both cyclic and pregnant endometrium and significantly higher in the endometrium at Day 35 of gestation than the cyclic endometrium. In Western blot analysis, an approximately 65 kDa band was detected in the endometrium, and approximately 51 kDa in the cultured bovine epithelial cells and BT-1 cells, respectively. Both in situ hybridization and immunohistochemistry data showed that EMMPRIN was primarily expressed in luminal and glandular epithelium with strong staining on Day 19 conceptus. At Day 19 of gestation, expression of EMMPRIN mRNA on luminal epithelium was decreased than that observed at middle of estrous cycle, however, on Day 30 of gestation, slightly increased expression was found at the site of placentation. Expression of matrix metalloproteinase-2 (MMP-2) and MMP-14 mRNA were mainly detected in stroma and their expression also decreased at Day 19 of gestation however it was also expressed at the site of placentation at Day 30 of gestation as observed for EMMPRIN. Expression of MMP-1 or -9 mRNA was very low and was below the detection limit in the cyclic and pregnant endometrium. Conclusion EMMPRIN from the luminal epithelium may regulate the expression of stromal MMP-2 and -14 suggesting its crucial role in

  11. Naturally Occurring Extracellular Matrix Scaffolds for Dermal Regeneration: Do They Really Need Cells?

    PubMed Central

    Eweida, A. M.; Marei, M. K.

    2015-01-01

    The pronounced effect of extracellular matrix (ECM) scaffolds in supporting tissue regeneration is related mainly to their maintained 3D structure and their bioactive components. These decellularized matrix scaffolds could be revitalized before grafting via adding stem cells, fibroblasts, or keratinocytes to promote wound healing. We reviewed the online published literature in the last five years for the studies that performed ECM revitalization and discussed the results of these studies and the related literature. Eighteen articles met the search criteria. Twelve studies included adding cells to acellular dermal matrix (ADM), 3 studies were on small intestinal mucosa (SIS), one study was on urinary bladder matrix (UBM), one study was on amniotic membrane, and one study included both SIS and ADM loaded constructs. We believe that, in chronic and difficult-to-heal wounds, revitalizing the ECM scaffolds would be beneficial to overcome the defective host tissue interaction. This belief still has to be verified by high quality randomised clinical trials, which are still lacking in literature. PMID:26509165

  12. Ubiquitylation Functions in the Calcium Carbonate Biomineralization in the Extracellular Matrix

    PubMed Central

    Fang, Dong; Pan, Cong; Lin, Huijuan; Lin, Ya; Xu, Guangrui; Zhang, Guiyou; Wang, Hongzhong; Xie, Liping; Zhang, Rongqing

    2012-01-01

    Mollusks shell formation is mediated by matrix proteins and many of these proteins have been identified and characterized. However, the mechanisms of protein control remain unknown. Here, we report the ubiquitylation of matrix proteins in the prismatic layer of the pearl oyster, Pinctada fucata. The presence of ubiquitylated proteins in the prismatic layer of the shell was detected with a combination of western blot and immunogold assays. The coupled ubiquitins were separated and identified by Edman degradation and liquid chromatography/mass spectrometry (LC/MS). Antibody injection in vivo resulted in large amounts of calcium carbonate randomly accumulating on the surface of the nacreous layer. These ubiquitylated proteins could bind to specific faces of calcite and aragonite, which are the two main mineral components of the shell. In the in vitro calcium carbonate crystallization assay, they could reduce the rate of calcium carbonate precipitation and induce the calcite formation. Furthermore, when the attached ubiquitins were removed, the functions of the EDTA-soluble matrix of the prismatic layer were changed. Their potency to inhibit precipitation of calcium carbonate was decreased and their influence on the morphology of calcium carbonate crystals was changed. Taken together, ubiquitylation is involved in shell formation. Although the ubiquitylation is supposed to be involved in every aspect of biophysical processes, our work connected the biomineralization-related proteins and the ubiquitylation mechanism in the extracellular matrix for the first time. This would promote our understanding of the shell biomineralization and the ubiquitylation processes. PMID:22558208

  13. The Extracellular Matrix Component Psl Provides Fast-Acting Antibiotic Defense in Pseudomonas aeruginosa Biofilms

    PubMed Central

    Billings, Nicole; Ramirez Millan, Maria; Caldara, Marina; Rusconi, Roberto; Tarasova, Yekaterina; Stocker, Roman; Ribbeck, Katharina

    2013-01-01

    Bacteria within biofilms secrete and surround themselves with an extracellular matrix, which serves as a first line of defense against antibiotic attack. Polysaccharides constitute major elements of the biofilm matrix and are implied in surface adhesion and biofilm organization, but their contributions to the resistance properties of biofilms remain largely elusive. Using a combination of static and continuous-flow biofilm experiments we show that Psl, one major polysaccharide in the Pseudomonas aeruginosa biofilm matrix, provides a generic first line of defense toward antibiotics with diverse biochemical properties during the initial stages of biofilm development. Furthermore, we show with mixed-strain experiments that antibiotic-sensitive “non-producing” cells lacking Psl can gain tolerance by integrating into Psl-containing biofilms. However, non-producers dilute the protective capacity of the matrix and hence, excessive incorporation can result in the collapse of resistance of the entire community. Our data also reveal that Psl mediated protection is extendible to E. coli and S. aureus in co-culture biofilms. Together, our study shows that Psl represents a critical first bottleneck to the antibiotic attack of a biofilm community early in biofilm development. PMID:23950711

  14. Quantification of fibronectin as a method to assess ex vivo extracellular matrix remodeling.

    PubMed

    Bager, C L; Gudmann, N; Willumsen, N; Leeming, D J; Karsdal, M A; Bay-Jensen, A C; Høgdall, E; Balslev, I; He, Y

    2016-09-16

    Altered architecture, composition and quality of the extracellular matrix (ECM) are pathological hallmarks of several inflammatory and fibro-proliferative pathological processes such as osteoarthritis (OA), rheumatoid arthritis (RA), fibrosis and cancer. One of the most important components of the ECM is fibronectin. Fibronectin serves as an adhesion molecule anchoring cells to the underlying basement membrane through direct interaction with integrin receptors. Fibronectin hereby modulates the properties of the ECM and affects cellular processes. Quantification of fibronectin remodeling could therefore be used to assess the changes in the ECM that occur during progression of fibro-proliferative pathologies. Ex vivo models are becoming state-of-the-art tools to study ECM remodeling as the cellular composition and the organization of the ECM are preserved. Ex vivo models may therefore be a valuable tool to study the ECM remodeling that occurs during progression of fibro-proliferative pathologies. The aim of this study was to quantify fibronectin remodeling in ex vivo models of cartilage and cancer. A competitive The enzyme-linked immunosorbent assay (ELISA) against the C-terminus of fibronectin was developed (FBN-C). The assay was evaluated in relation to specificity, technical performance and as a marker for quantification of fibronectin in cartilage and cancer ex vivo models. The ELISA was specific and technically stable. Cleavage of tumor tissue with MMP-2 released significantly higher levels of FBN-C compared to tissue with buffer only and western blot analysis revealed that FBN-C recognizes both full length and degraded fibronectin. When ex vivo cartilage cultures were stimulated with the anabolic factor TGFβ and catabolic factors TNF-α and OSM, significantly higher levels of FBN-C were found in the conditioned media. Lastly, FBN-C was released from a cancer ex vivo model. In conclusion, we were able to quantify fibronectin remodeling in ex vivo models

  15. The Notch pathway mediates the angiotensin II-induced synthesis of extracellular matrix components in podocytes.

    PubMed

    Yao, Min; Wang, Xiaomei; Wang, Xiaomeng; Zhang, Tao; Chi, Yanqing; Gao, Feng

    2015-07-01

    The Notch pathway is known to contribute to the development of glomerular disease. Angiotensin II (Ang II), an important member of the renin-angiotensin system, stimulates the accumulation of extracellular matrix components in glomerular disease; however, the exact mechanisms involved remain to be elucidated. In the present study, we aimed to investigate the effects of the Notch pathway on the synthesis of extracellular matrix components in Ang II-stimulated podocytes. Mouse podocytes were stimulated with Ang II (10-6 mol/l). The activation of the Notch pathway was inhibited by a vector carrying short hairpin RNA (shRNA) targeting Notch1 (sh-Notch1) or by γ-secretase inhibitor (GSI). The protein levels of Notch1, Notch intracellular domain 1 (NICD1), hairy and enhancer of split-1 (Hes1), matrix metalloproteinase (MMP)-2, MMP-9, transforming growth factor-β1 (TGF-β1), type IV collagen and laminin were determined by western blot analysis. The Notch1, Hes1, MMP-2, MMP-9, TGF-β1, type IV collagen and laminin mRNA levels were detected by RT-PCR. The MMP-2 and MMP-9 activity was measured using a cell active fluorescence assay kit. The levels of TGF-β1, type IV collagen and laminin were determined in the culture medium of the podocytes by enzyme-linked immunosorbent assay (ELISA). Our results revealed that Ang II upregulated Notch1, NICD1, Hes1, TGF-β1, type IV collagen and laminin expression and downregulated MMP-2 and MMP-9 expression in the cultured podocytes. The inhibition of the Notch pathway by sh-Notch1 or GSI increased MMP-2 and MMP-9 expression, decreased the TGF-β1 level and suppressed type IV collagen and laminin expression. The inhibition of the Notch pathway by sh-Notch1 or GSI also increased MMP-2 and MMP-9 activity, and decreased TGF-β1 levels, type IV collagen levels and laminin secretion. These findings indicate that the Notch pathway potentially mediates the Ang II-induced synthesis of extracellular matrix components in podocytes through the

  16. Mechanisms of cytoskeletal regulation. Modulation of aortic endothelial cell spectrin by the extracellular matrix.

    PubMed Central

    Pratt, B. M.; Harris, A. S.; Morrow, J. S.; Madri, J. A.

    1984-01-01

    Endothelial cells have a complex cytoskeleton that is responsive to a variety of stimuli such as shear and desquamative injury. The extracellular matrix is known to influence several aspects of cellular behavior, including attachment, spreading, and migration and may, in part, initiate and control vascular responses in growth, differentiation, wound repair, and neoplasia. It is likely that linkage between surface receptors responsible for sensing the matrix and the cytoskeleton may be relevant to understanding the mechanisms of these responses. Spectrin is a high-molecular-weight heterodimer recently identified in many cells that appears to link surface receptors to cortical actin filaments. We have confirmed the existence of spectrin in cultured aortic endothelial cells by metabolic labeling and immunoprecipitation and demonstrated that its organization and intracellular distribution is sensitive to the extracellular matrix. When bovine calf aortic endothelial cells (BAEC) are cultured to confluency on a fibronectin (Fn) substrate, they assume a flattened, spread morphology and exhibit a punctate spectrin distribution with no discernible peripheral localization. In contrast, BAECs cultured on a Type I/III collagen (I/III) substrate exhibit a fibrillar spectrin pattern with significant peripheral localization. When migrating cells were examined, the distribution of spectrin was strikingly different. The cells on the Fn substrate showed no changes in spectrin localization, whereas the cells on I/III exhibited a significant rearrangement, with spectrin being in a coarse fibrillar form, with the fibrils aligned parallel to the direction of migration. The differences in arrangement of this cytoskeletal component on the two substrata reflect the ability of the substrate to perturb the cytoskeletal organization and modulate some aspects of cell behavior such as spreading, proliferation, and migration. These data are consistent with the concept that the nonerythroid

  17. Neutrophil elastase processing of Gelatinase A is mediated by extracellular matrix

    SciTech Connect

    Rice, A.; Banda, M.J.

    1995-07-18

    Gelatinase A (72-kDa type IV collagenase) is a metalloproteinase that is expressed by many cells in culture and is overexpressed by some tumor cells. It has been suggested that the serine proteinase neutrophil elastase might play a role iii the posttranslational processing of gelatinase A and that noncatalytic interactions between gelatinase A and components of the extracellular matrix might alter potential processing pathways. These questions were addressed with the use of gelatin substrate zymography, gelatinolytic activity assays, and amino acid sequence analysis. We found that neutrophil elastase does proteolytically modify gelatinase A by cleaving at a number of sites within gelatinase A. Sequential treatment of gelatinase A with 4-aminophenylmercuric acetate (APMA) and neutrophil elastase yielded an active gelatinase with a 4-fold increase in gelatinolytic activity. The increased gelatinolytic activity correlated with that of a 40-kDa fragment of gelatinase A. Matrix components altered the proteolytic modifications in gelatinase A that were mediated by neutrophil elastase. In the absence of gelatin, neutrophil elastase destructively degraded gelatinase A by hydrolyzing at least two bonds within the fibronectin-like gelatin-binding domain of gelatinase A. In the presence of gelatin, these two inactivating cleavage sites were protected, and cleavage at a site within the hemopexin-like carboxyl-terminal domain resulted in a truncated yet active gelatinase. The results suggest a regulatory role for extracellular matrix molecules in stabilizing gelatinase A fragments and in altering the availability of sites susceptible to destructive proteolysis by neutrophil elastase. 32 refs., 10 figs.

  18. Biological functionality of extracellular matrix-ornamented three-dimensional printed hydroxyapatite scaffolds.

    PubMed

    Kumar, A; Nune, K C; Misra, R D K

    2016-06-01

    Three-dimensional (3D) printing is considered an ideally suitable method to fabricate patient specific implantable devices. The approach enabled to produce a porous scaffold with tailored physical, mechanical, and biological properties because of the flexibility to tune the scaffold architecture. The objective of the study described was to elucidate the determining role of cell-laid extracellular matrix (ECM) in impacting biological response. In this regard, to mimic the natural ECM environment or the attributes of the native tissue, a natural ECM analogue surface was produced on the 3D printed and sintered hydroxyapatite (HA) scaffold surface by the mineralized ECM of the osteoblast. This involved the growth of osteoblast on 3D printed scaffolds, followed by differentiation to deposit the mineralized ECM on the biomaterial surface. The cells were removed from the mineralized matrix using freeze-thaw cycles to obtain a decellularized extracellular matrix (dECM) on the biomaterial surface. Subsequently, seeding of osteoblast on dECM-ornamented HA scaffolds led to 3D growth with enhanced expression of prominent proteins, actin and vinculin. Based on preliminary observations of present study, it was underscored that HA scaffolds-ornamented with dECM provided an optimized microenvironment conducive to the growth of 3D structural tissue and favorably promoted biological functionality because of the availability of an environment that promoted cell-cell and cell-scaffold interaction. The primary advantage of dECM is that it enabled constructive remodeling and promoted the formation of tissue in lieu of less functional tissue. The study opens-up a new path for printing of 3D structures suitable to treat segmental bone defects. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1343-1351, 2016. PMID:26799466

  19. Growth and morphogenesis of embryonic mouse organs on non-coated and extracellular matrix-coated Biopore membrane

    NASA Technical Reports Server (NTRS)

    Hardman, P.; Klement, B. J.; Spooner, B. S.

    1993-01-01

    Embryonic mouse salivary glands, pancreata, and kidneys were isolated from embryos of appropriate gestational age by microdissection, and were cultured on Biopore membrane either non-coated or coated with type I collagen or Matrigel. As expected, use of Biopore membrane allowed high quality photomicroscopy of the living organs. In all organs extensive mesenchymal spreading was observed in the presence of type I collagen or Matrigel. However, differences were noted in the effects of extracellular matrix (ECM) coatings on epithelial growth and morphogenesis: salivary glands were minimally affected, pancreas morphogenesis was adversely affected, and kidney growth and branching apparently was enhanced. It is suggested that these differences in behaviour reflect differences in the strength of interactions between the mesenchymal cells and their surrounding endogenous matrix, compared to the exogenous ECM macromolecules. This method will be useful for culture of these and other embryonic organs. In particular, culture of kidney rudiments on ECM-coated Biopore offers a great improvement over previously used methods which do not allow morphogenesis to be followed in vitro.

  20. Extracellular matrix from human umbilical cord-derived mesenchymal stem cells as a scaffold for peripheral nerve regeneration

    PubMed Central

    Xiao, Bo; Rao, Feng; Guo, Zhi-yuan; Sun, Xun; Wang, Yi-guo; Liu, Shu-yun; Wang, Ai-yuan; Guo, Quan-yi; Meng, Hao-ye; Zhao, Qing; Peng, Jiang; Wang, Yu; Lu, Shi-bi

    2016-01-01

    The extracellular matrix, which includes collagens, laminin, or fibronectin, plays an important role in peripheral nerve regeneration. Recently, a Schwann cell-derived extracellular matrix with classical biomaterial was used to mimic the neural niche. However, extensive clinical use of Schwann cells remains limited because of the limited origin, loss of an autologous nerve, and extended in vitro culture times. In the present study, human umbilical cord-derived mesenchymal stem cells (hUCMSCs), which are easily accessible and more proliferative than Schwann cells, were used to prepare an extracellular matrix. We identified the morphology and function of hUCMSCs and investigated their effect on peripheral nerve regeneration. Compared with a non-coated dish tissue culture, the hUCMSC-derived extracellular matrix enhanced Schwann cell proliferation, upregulated gene and protein expression levels of brain-derived neurotrophic factor, glial cell-derived neurotrophic factor, and vascular endothelial growth factor in Schwann cells, and enhanced neurite outgrowth from dorsal root ganglion neurons. These findings suggest that the hUCMSC-derived extracellular matrix promotes peripheral nerve repair and can be used as a basis for the rational design of engineered neural niches.

  1. Shotgun proteomics implicates extracellular matrix proteins and protease systems in neuronal development induced by astrocyte cholinergic stimulation

    PubMed Central

    Moore, Nadia H.; Costa, Lucio G.; Shaffer, Scott A; Goodlett, David R.; Guizzetti, Marina

    2009-01-01

    Astrocytes play an important role in neuronal development through the release of soluble factors that affect neuronal maturation. Shotgun proteomics followed by Gene Ontology analysis was used in this study to identify proteins present in the conditioned medium of primary rat astrocytes. 133 secreted proteins were identified, the majority of which were never before reported to be produced by astrocytes. Extracellular proteins were classified based on their biological and molecular functions; most of the identified proteins were involved in neuronal development. Semi-quantitative proteomic analysis was carried out to identify changes in the levels of proteins released by astrocytes after stimulation with the cholinergic agonist carbachol, as we have previously reported that carbachol-treated astrocytes elicit neuritogenesis in hippocampal neurons through the release of soluble factors. Carbachol up-regulated the secretion of 15 proteins and down-regulated the release of 17 proteins. Changes in the levels of four proteins involved in neuronal differentiation (thrombospondin-1, fibronectin, plasminogen activator inhibitor-1, and plasminogen activator urokinase) were verified by Western blot or ELISA. In conclusion, this study identified a large number of proteins involved in neuronal development in the astrocyte secretome and implicated extracellular matrix proteins and protease systems in neuronal development induced by astrocyte cholinergic stimulation. PMID:19077055

  2. FAP-overexpressing fibroblasts produce an extracellular matrix that enhances invasive velocity and directionality of pancreatic cancer cells

    PubMed Central

    2011-01-01

    Background Alterations towards a permissive stromal microenvironment provide important cues for tumor growth, invasion, and metastasis. In this study, Fibroblast activation protein (FAP), a serine protease selectively produced by tumor-associated fibroblasts in over 90% of epithelial tumors, was used as a platform for studying tumor-stromal interactions. We tested the hypothesis that FAP enzymatic activity locally modifies stromal ECM (extracellular matrix) components thus facilitating the formation of a permissive microenvironment promoting tumor invasion in human pancreatic cancer. Methods We generated a tetracycline-inducible FAP overexpressing fibroblastic cell line to synthesize an in vivo-like 3-dimensional (3D) matrix system which was utilized as a stromal landscape for studying matrix-induced cancer cell behaviors. A FAP-dependent topographical and compositional alteration of the ECM was characterized by measuring the relative orientation angles of fibronectin fibers and by Western blot analyses. The role of FAP in the matrix-induced permissive tumor behavior was assessed in Panc-1 cells in assorted matrices by time-lapse acquisition assays. Also, FAP+ matrix-induced regulatory molecules in cancer cells were determined by Western blot analyses. Results We observed that FAP remodels the ECM through modulating protein levels, as well as through increasing levels of fibronectin and collagen fiber organization. FAP-dependent architectural/compositional alterations of the ECM promote tumor invasion along characteristic parallel fiber orientations, as demonstrated by enhanced directionality and velocity of pancreatic cancer cells on FAP+ matrices. This phenotype can be reversed by inhibition of FAP enzymatic activity during matrix production resulting in the disorganization of the ECM and impeded tumor invasion. We also report that the FAP+ matrix-induced tumor invasion phenotype is β1-integrin/FAK mediated. Conclusion Cancer cell invasiveness can be affected by

  3. The extracellular matrix of hydra is a porous sheet and contains type IV collagen.

    PubMed

    Shimizu, Hiroshi; Aufschnaiter, Roland; Li, Li; Sarras, Michael P; Borza, Dorin-Bogdan; Abrahamson, Dale R; Sado, Yoshikazu; Zhang, Xiaoming

    2008-01-01

    Hydra, as an early diploblastic metazoan, has a well-defined extracellular matrix (ECM) called mesoglea. It is organized in a tri-laminar pattern with one centrally located interstitial matrix that contains type I collagen and two sub-epithelial zones that resemble a basal lamina containing laminin and possibly type IV collagen. This study used monoclonal antibodies to the three hydra mesoglea components (type I, type IV collagens and laminin) and immunofluorescent staining to visualize hydra mesoglea structure and the relationship between these mesoglea components. In addition, hydra mesoglea was isolated free of cells and studied with immunofluorescence and scanning electron microscopy (SEM). Our results show that type IV collagen co-localizes with laminin in the basal lamina whereas type I collagen forms a grid pattern of fibers in the interstitial matrix. The isolated mesoglea can maintain its structural stability without epithelial cell attachment. Hydra mesoglea is porous with multiple trans-mesoglea pores ranging from 0.5 to 1 microm in diameter and about six pores per 100 microm(2) in density. We think these trans-mesoglea pores provide a structural base for epithelial cells on both sides to form multiple trans-mesoglea cell-cell contacts. Based on these findings, we propose a new model of hydra mesoglea structure. PMID:18602803

  4. Mechanical forces regulate the interactions of fibronectin and collagen I in extracellular matrix

    PubMed Central

    Kubow, Kristopher E.; Vukmirovic, Radmila; Zhe, Lin; Klotzsch, Enrico; Smith, Michael L.; Gourdon, Delphine; Luna, Sheila; Vogel, Viola

    2015-01-01

    Despite the crucial role of extracellular matrix (ECM) in directing cell fate in healthy and diseased tissues—particularly in development, wound healing, tissue regeneration and cancer—the mechanisms that direct the assembly and regulate hierarchical architectures of ECM are poorly understood. Collagen I matrix assembly in vivo requires active fibronectin (Fn) fibrillogenesis by cells. Here we exploit Fn-FRET probes as mechanical strain sensors and demonstrate that collagen I fibres preferentially co-localize with more-relaxed Fn fibrils in the ECM of fibroblasts in cell culture. Fibre stretch-assay studies reveal that collagen I's Fn-binding domain is responsible for the mechano-regulated interaction. Furthermore, we show that Fn-collagen interactions are reciprocal: relaxed Fn fibrils act as multivalent templates for collagen assembly, but once assembled, collagen fibres shield Fn fibres from being stretched by cellular traction forces. Thus, in addition to the well-recognized, force-regulated, cell-matrix interactions, forces also tune the interactions between different structural ECM components. PMID:26272817

  5. Extracellular matrix biomimicry for the creation of investigational and therapeutic devices.

    PubMed

    Pellowe, Amanda S; Gonzalez, Anjelica L

    2016-01-01

    The extracellular matrix (ECM) is a web of fibrous proteins that serves as a scaffold for tissues and organs, and is important for maintaining homeostasis and facilitating cellular adhesion. Integrin transmembrane receptors are the primary adhesion molecules that anchor cells to the ECM, thus integrating cells with their microenvironments. Integrins play a critical role in facilitating cell-matrix interactions and promoting signal transduction, both from the cell to the ECM and vice versa, ultimately mediating cell behavior. For this reason, many advanced biomaterials employ biomimicry by replicating the form and function of fibrous ECM proteins. The ECM also acts as a reservoir for small molecules and growth factors, wherein fibrous proteins directly bind and present these bioactive moieties that facilitate cell activity. Therefore biomimicry can be enhanced by incorporating small molecules into ECM-like substrates. Biomimetic ECM materials have served as invaluable research tools for studying interactions between cells and the surrounding ECM, revealing that cell-matrix signaling is driven by mechanical forces, integrin engagement, and small molecules. Mimicking pathological ECMs has also elucidated disease specific cell behaviors. For example, biomimetic tumor microenvironments have been used to induce metastatic cell behaviors, and have thereby shown promise for in vitro cancer drug testing and targeting. Further, ECM-like substrates have been successfully employed for autologous cell recolonization for tissue engineering and wound healing. As we continue to learn more about the mechanical and biochemical characteristics of the ECM, these properties can be harnessed to develop new biomaterials, biomedical devices, and therapeutics. PMID:26053111

  6. Analysis of adherence of Streptococcus defectivus and endocarditis-associated streptococci to extracellular matrix.

    PubMed Central

    Tart, R C; van de Rijn, I

    1991-01-01

    Pathogenesis of nutritionally variant streptococcal (NVS) endocarditis initiates with bacterial attachment to and colonization of the damaged heart valve surface. Underlying extracellular matrix (ECM) exposed to the environment during damage to cardiac endothelium provides additional receptors that could be involved in bacterial adherence. The ability of NVS and endocarditis-associated streptococci to bind ECM was investigated by using an enzyme-linked immunosorbent assay system that incorporated ECM secreted by baby hamster kidney and human umbilical vein endothelial cells in culture. Streptococcus defectivus, the major species isolated from NVS endocarditis cases, bound ECM of fibroblasts and endothelial cells, indicating that the ECM molecule involved in the binding was a common constituent of diverse matrices. The specific binding of S. defectivus to ECM was demonstrated by saturation binding and specific antibody inhibition studies. Of the 15 S. defectivus strains analyzed, 13 bound ECM, whereas Streptococcus adjacens and NVS serotype III strains were unable to bind the matrix. This selective binding suggested that S. defectivus binds to heart valves through a mechanism different from those of other NVS in subacute bacterial endocarditis. A survey of non-NVS streptococcal endocarditis isolates demonstrated that S. mutans, S. mitis, S. sanguis, and S. faecalis also bound ECM, whereas other viridans species were unable to bind the matrix. Images PMID:1997435

  7. The extracellular matrix of hydra is a porous sheet and contains type IV collagen

    PubMed Central

    Shimizu, Hiroshi; Aufschnaiter, Roland; Li, Li; Sarras, Michael P.; Borza, Dorin-Bogdan; Abrahamson, Dale R.; Sado, Yoshikazu; Zhang, Xiaoming

    2008-01-01

    Hydra, as an early diploblastic metazoan, has a well defined extracellular matrix (ECM) called-mesoglea. It is organized in a tri-laminar pattern with one centrally located interstitial matrix that contains type I collagen and two sub-epithelial zones that resemble a basal lamina containing laminin and possibly type IV collagen. This study used monoclonal antibodies to the three hydra mesoglea components (type I, type IV collagens and laminin) and immunofluorescent staining to visualize hydra mesoglea structure and the relationship between these mesoglea components. In addition, hydra mesoglea was isolated free of cells and studied with immunofluorescence and SEM. Our results show that type IV collagen co-localizes with laminin in the basal lamina whereas type I collagen forms a grid pattern of fibers in the interstitial matrix. The isolated-mesoglea can maintain its structural stability without epithelial cell attachment. Hydra mesogleais porous with multiple trans-mesoglea pores ranging from 0.5 to 1 µm in diameter and about 6 pores per 100 µm2 in density. We think these trans-mesoglea pores provide a structural base for epithelial cells on both sides to form multiple trans-mesoglea cell-cell contacts. Based on these findings, we propose a new model of hydra mesoglea structure. PMID:18602803

  8. Early alterations in extracellular matrix and transforming growth factor [beta] gene expression in mouse lung indicative of late radiation fibrosis

    SciTech Connect

    Finkelstein, J.N.; Johnston, C.J.; Baggs, R.; Rubin, P. )

    1994-02-01

    Fibrosis, characterized by the accumulation of collagen, is a late result of thoracic irradiation. The expression of late radiation injury can be found immediately after irradiation by measuring messenger RNA (mRNA) abundance. To determine if extracellular matrix mRNA and transforming growth factor beta abundance was affected acutely after irradiation, the authors measured mRNA levels of collagen I (CI), collagen III (CIII), collagen IV (CIV), fibronectin (FN), and transforming growth factor [beta] (TGF[beta][sub 1,2 3]) in mouse lungs on day 1 and day 14 after graded doses of radiation. C57BL/6 female mice were irradiated with a single dose to the thorax of 5 or 12.5 Gy. Total lung RNA was prepared and immobilized by Northern and slot blotting and hybridized with radiolabelled cDNA probes for CI, CIII, CIV, FN, TGF[beta][sub 1,2 3] and a control probe encoding for glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Autoradiographic data were quantified by video densitometry and results normalized to GAPDH. Changes in the expression of CI, CIII, CIV, FN and TGF[beta][sub 1,2 3] were observed as early as 1 day after exposure. Through 14 days, changes in mRNA up to 5-fold were seen for any one dose. Dose related changes as high as 10-fold were also evident. The CI:CIII ratio increased gradually for the 5 Gy dose at 14 days postirradiation while the CI:CII ratio for the 12.5 Gy dose decreased by approximately 4-fold as compared to the control. These studies suggest that alterations in expression of extracellular matrix and TGF[beta] mRNA occur very early after radiation injury even at low doses and may play a role in the development of chronic fibrosis. 37 refs., 6 figs.

  9. The pherophorins: common, versatile building blocks in the evolution of extracellular matrix architecture in Volvocales.

    PubMed

    Hallmann, Armin

    2006-01-01

    Green algae of the order Volvocales provide an unrivalled opportunity for exploring the transition from unicellularity to multicellularity. They range from unicells, like Chlamydomonas, through homocytic colonial forms with increasing cooperation of individual cells, like Gonium or Pandorina, to heterocytic multicellular forms with different cell types and a complete division of labour, like Volvox. A fundamental requirement for the evolution of multicellularity is the development of a complex, multifunctional extracellular matrix (ECM). The ECM has many functions, which can change under developmental control or as a result of environmental factors. Here molecular data from 15 novel proteins are presented. These proteins have been identified in Chlamydomonas reinhardtii, Gonium pectorale, Pandorina morum and Volvox carteri, and all belong to a single protein family, the pherophorins. Pherophorin-V1 is shown to be a glycoprotein localized to the 'cellular zone' of the V. carteri ECM. Pherophorin-V1 and -V2 mRNAs are strongly induced not only by the sex inducer, which triggers sexual development at extremely low concentrations, but also by mechanical wounding. Like the extensins of higher plants, which are also developmentally controlled or sometimes inducible by wounding, the pherophorins contain a (hydroxy-)proline-rich (HR) rod-like domain and are abundant within the extracellular compartment. In contrast to most extensins, pherophorins have additional globular A and B domains on both ends of the HR domains. Therefore pherophorins most closely resemble a particular class of higher plant extensin, the solanaceous lectins (e.g. potato lectin), suggesting multivalent carbohydrate-binding functions are present within the A and B domains and are responsible for cross-linking. Our results suggest that pherophorins are used as the building blocks for the extracellular scaffold throughout the Volvocales, with the characteristic mesh sizes in different ECM structures being

  10. Prohibitin Attenuates Oxidative Stress and Extracellular Matrix Accumulation in Renal Interstitial Fibrosis Disease

    PubMed Central

    Qin, Yuan-Han; Lei, Feng-Ying; Huang, Wei-Fang

    2013-01-01

    Prohibitin is an evolutionary conserved and pleiotropic protein that has been implicated in various cellular functions, including proliferation, tumour suppression, apoptosis, transcription, and mitochondrial protein folding. Both prohibitin over- and under-expression have been implicated in various diseases and cell types. We recently demonstrated that prohibitin down-regulation results in increased renal interstitial fibrosis (RIF). Here we investigated the role of oxidative stress and prohibitin expression in RIF in unilateral ureteral obstructed rats. Lentivirus-based delivery vectors were used to knockdown or over-express prohibitin. Our results show that increased prohibitin expression was negatively correlated with the RIF index, reactive oxygen species, malon dialdehyde, transforming growth factor β1, collagen IV, fibronectin, and cell apoptosis index. In conclusion, we postulate that prohibitin acts as a positive regulator of mechanisms that counteract oxidative stress and extracellular matrix accumulation and therefore has an antioxidative effect. PMID:24204768

  11. Extracellular matrix of secondary lymphoid organs impacts on B-cell fate and survival

    PubMed Central

    Horn, Nathalie; Ruegg, Markus A.; Sonnenberg, Arnoud; Georges-Labouesse, Elisabeth; Winkler, Thomas H.; Kearney, John F.; Cardell, Susanna; Sorokin, Lydia

    2013-01-01

    We describe a unique extracellular matrix (ECM) niche in the spleen, the marginal zone (MZ), characterized by the basement membrane glycoproteins, laminin α5 and agrin, that promotes formation of a specialized population of MZ B lymphocytes that respond rapidly to blood-borne antigens. Mice with reduced laminin α5 expression show reduced MZ B cells and increased numbers of newly formed (NF) transitional B cells that migrate from the bone marrow, without changes in other immune or stromal cell compartments. Transient integrin α6β1-mediated interaction of NF B cells with laminin α5 in the MZ supports the MZ B-cell population, their long-term survival, and antibody response. Data suggest that the unique 3D structure and biochemical composition of the ECM of lymphoid organs impacts on immune cell fate. PMID:23847204

  12. Three-Dimensional Characterization of Mechanical Interactions between Endothelial Cells and Extracellular Matrix during Angiogenic Sprouting

    PubMed Central

    Du, Yue; Herath, Sahan C. B.; Wang, Qing-guo; Wang, Dong-an; Asada, H. Harry; Chen, Peter C. Y.

    2016-01-01

    We studied the three-dimensional cell-extracellular matrix interactions of endothelial cells that form multicellular structures called sprouts. We analyzed the data collected in-situ from angiogenic sprouting experiments and identified the differentiated interaction behavior exhibited by the tip and stalk cells. Moreover, our analysis of the tip cell lamellipodia revealed the diversity in their interaction behavior under certain conditions (e.g., when the heading of a sprout is switched approximately between the long-axis direction of two different lamellipodia). This study marks the first time that new characteristics of such interactions have been identified with shape changes in the sprouts and the associated rearrangements of collagen fibers. Clear illustrations of such changes are depicted in three-dimensional views. PMID:26903154

  13. Biomaterials approaches to modeling macrophage-extracellular matrix interactions in the tumor microenvironment.

    PubMed

    Springer, Nora L; Fischbach, Claudia

    2016-08-01

    Tumors are characterized by aberrant extracellular matrix (ECM) remodeling and chronic inflammation. While advances in biomaterials and tissue engineering strategies have led to important new insights regarding the role of ECM composition, structure, and mechanical properties in cancer in general, the functional link between these parameters and macrophage phenotype is poorly understood. Nevertheless, increasing experimental evidence suggests that macrophage behavior is similarly controlled by physicochemical properties of the ECM and consequential changes in mechanosignaling. Here, we will summarize the current knowledge of macrophage biology and ECM-mediated differences in mechanotransduction and discuss future opportunities of biomaterials and tissue engineering platforms to interrogate the functional relationship between these parameters and their relevance to cancer. PMID:26921768

  14. Patterned Anchorage to the Apical Extracellular Matrix Defines Tissue Shape in the Developing Appendages of Drosophila

    PubMed Central

    Ray, Robert P.; Matamoro-Vidal, Alexis; Ribeiro, Paulo S.; Tapon, Nic; Houle, David; Salazar-Ciudad, Isaac; Thompson, Barry J.

    2015-01-01

    Summary How tissues acquire their characteristic shape is a fundamental unresolved question in biology. While genes have been characterized that control local mechanical forces to elongate epithelial tissues, genes controlling global forces in epithelia have yet to be identified. Here, we describe a genetic pathway that shapes appendages in Drosophila by defining the pattern of global tensile forces in the tissue. In the appendages, shape arises from tension generated by cell constriction and localized anchorage of the epithelium to the cuticle via the apical extracellular-matrix protein Dumpy (Dp). Altering Dp expression in the developing wing results in predictable changes in wing shape that can be simulated by a computational model that incorporates only tissue contraction and localized anchorage. Three other wing shape genes, narrow, tapered, and lanceolate, encode components of a pathway that modulates Dp distribution in the wing to refine the global force pattern and thus wing shape. PMID:26190146

  15. Bromelain surface modification increases the diffusion of silica nanoparticles in the tumor extracellular matrix.

    PubMed

    Parodi, Alessandro; Haddix, Seth G; Taghipour, Nima; Scaria, Shilpa; Taraballi, Francesca; Cevenini, Armando; Yazdi, Iman K; Corbo, Claudia; Palomba, Roberto; Khaled, Sm Z; Martinez, Jonathan O; Brown, Brandon S; Isenhart, Lucas; Tasciotti, Ennio

    2014-10-28

    Tumor extracellular matrix (ECM) represents a major obstacle to the diffusion of therapeutics and drug delivery systems in cancer parenchyma. This biological barrier limits the efficacy of promising therapeutic approaches including the delivery of siRNA or agents intended for thermoablation. After extravasation due to the enhanced penetration and retention effect of tumor vasculature, typical nanotherapeutics are unable to reach the nonvascularized and anoxic regions deep within cancer parenchyma. Here, we developed a simple method to provide mesoporous silica nanoparticles (MSN) with a proteolytic surface. To this extent, we chose to conjugate MSN to Bromelain (Br-MSN), a crude enzymatic complex, purified from pineapple stems, that belongs to the peptidase papain family. This surface modification increased particle uptake in endothelial, macrophage, and cancer cell lines with minimal impact on cellular viability. Most importantly Br-MSN showed an increased ability to digest and diffuse in tumor ECM in vitro and in vivo. PMID:25119793

  16. Haemodynamic and extracellular matrix cues regulate the mechanical phenotype and stiffness of aortic endothelial cells

    PubMed Central

    Collins, Caitlin; Osborne, Lukas D.; Guilluy, Christophe; Chen, Zhongming; O’Brien, E Tim; Reader, John S.; Burridge, Keith; Superfine, Richard; Tzima, Ellie

    2014-01-01

    Endothelial cell (ECs) lining blood vessels express many mechanosensors, including platelet endothelial cell adhesion molecule-1 (PECAM-1), that convert mechanical force to biochemical signals. While it is accepted that mechanical stresses and the mechanical properties of ECs regulate vessel health, the relationship between force and biological response remains elusive. Here we show that ECs integrate mechanical forces and extracellular matrix (ECM) cues to modulate their own mechanical properties. We demonstrate that the ECM influences EC response to tension on PECAM-1. ECs adherent on collagen display divergent stiffening and focal adhesion growth compared to ECs on fibronectin. This is due to PKA-dependent serine phosphorylation and inactivation of RhoA. PKA signaling regulates focal adhesion dynamics and EC compliance in response to shear stress in vitro and in vivo. Our study identifies a ECM-specific, mechanosensitive signaling pathway that regulates EC compliance and may serve as an atheroprotective mechanism maintains blood vessel integrity in vivo. PMID:24917553

  17. Decellularized extracellular matrix microparticles as a vehicle for cellular delivery in a model of anastomosis healing.

    PubMed

    Hoganson, David M; Owens, Gwen E; Meppelink, Amanda M; Bassett, Erik K; Bowley, Chris M; Hinkel, Cameron J; Finkelstein, Eric B; Goldman, Scott M; Vacanti, Joseph P

    2016-07-01

    Extracellular matrix (ECM) materials from animal and human sources have become important materials for soft tissue repair. Microparticles of ECM materials have increased surface area and exposed binding sites compared to sheet materials. Decellularized porcine peritoneum was mechanically dissociated into 200 µm microparticles, seeded with fibroblasts and cultured in a low gravity rotating bioreactor. The cells avidly attached and maintained excellent viability on the microparticles. When the seeded microparticles were placed in a collagen gel, the cells quickly migrated off the microparticles and through the gel. Cells from seeded microparticles migrated to and across an in vitro anastomosis model, increasing the tensile strength of the model. Cell seeded microparticles of ECM material have potential for paracrine and cellular delivery therapies when delivered in a gel carrier. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1728-1735, 2016. PMID:26946064

  18. Crosslinking strategies for preparation of extracellular matrix-derived cardiovascular scaffolds

    PubMed Central

    Ma, Bing; Wang, Xiaoya; Wu, Chengtie; Chang, Jiang

    2014-01-01

    Heart valve and blood vessel replacement using artificial prostheses is an effective strategy for the treatment of cardiovascular disease at terminal stage. Natural extracellular matrix (ECM)-derived materials (decellularized allogeneic or xenogenic tissues) have received extensive attention as the cardiovascular scaffold. However, the bioprosthetic grafts usually far less durable and undergo calcification and progressive structural deterioration. Glutaraldehyde (GA) is a commonly used crosslinking agent for improving biocompatibility and durability of the natural scaffold materials. However, the nature ECM and GA-crosslinked materials may result in calcification and eventually lead to the transplant failure. Therefore, studies have been conducted to explore new crosslinking agents. In this review, we mainly focused on research progress of ECM-derived cardiovascular scaffolds and their crosslinking strategies. PMID:26816627

  19. NG2 proteoglycan increases mesangial cell proliferation and extracellular matrix production

    SciTech Connect

    Xiong Jing; Wang Yang; Zhu, Zhonghua; Liu Jianshe; Wang Yumei; Zhang Chun; Hammes, Hans-Peter; Lang, Florian; Feng Yuxi

    2007-10-05

    As a membrane-spanning protein, NG2 chondroitin sulfate proteoglycan interacts with molecules on both sides of plasma membrane. The present study explored the role of NG2 in the pathogenesis of diabetic nephropathy. In the normal kidneys, NG2 was observed predominantly in glomerular mesangium, Bowman's capsule and interstitial vessels. Both mRNA and protein expression in kidneys was significantly higher in strepozotocin-induced diabetic rats than that in normal rats. In the cultured rat mesangial cell line HBZY-1, overexpression of NG2 promoted mesangial cell proliferation and extracellular matrix (ECM) production, such as type VI collagen and laminin. Furthermore, target knockdown of NG2 resulted in decreased cell proliferation and ECM formation. The observations suggest that NG2 is up-regulated in diabetic nephropathy. It actively participates in the development and progression of glomerulosclerosis by stimulating proliferation of mesangial cells and deposition of ECM.

  20. Cell-mediated fibre recruitment drives extracellular matrix mechanosensing in engineered fibrillar microenvironments

    NASA Astrophysics Data System (ADS)

    Baker, Brendon M.; Trappmann, Britta; Wang, William Y.; Sakar, Mahmut S.; Kim, Iris L.; Shenoy, Vivek B.; Burdick, Jason A.; Chen, Christopher S.

    2015-12-01

    To investigate how cells sense stiffness in settings structurally similar to native extracellular matrices, we designed a synthetic fibrous material with tunable mechanics and user-defined architecture. In contrast to flat hydrogel surfaces, these fibrous materials recapitulated cell-matrix interactions observed with collagen matrices including stellate cell morphologies, cell-mediated realignment of fibres, and bulk contraction of the material. Increasing the stiffness of flat hydrogel surfaces induced mesenchymal stem cell spreading and proliferation; however, increasing fibre stiffness instead suppressed spreading and proliferation for certain network architectures. Lower fibre stiffness permitted active cellular forces to recruit nearby fibres, dynamically increasing ligand density at the cell surface and promoting the formation of focal adhesions and related signalling. These studies demonstrate a departure from the well-described relationship between material stiffness and spreading established with hydrogel surfaces, and introduce fibre recruitment as a previously undescribed mechanism by which cells probe and respond to mechanics in fibrillar matrices.

  1. Isolation and culture of human trabecular meshwork cells by extracellular matrix digestion.

    PubMed

    Stamer, W D; Seftor, R E; Williams, S K; Samaha, H A; Snyder, R W

    1995-07-01

    Like corneal endothelial cells, human trabecular meshwork cells are believed to be of neural crest origin, but demonstrate physiological properties and an antithrombogenic surface similar to vascular endothelial cells. One current method for isolating trabecular meshwork cells utilizes the motile nature of these cells to migrate away from a trabecular meshwork explant in culture to more distal regions of the culture dish. This 'outgrowth' technique is limited in practice by the relatively small number of cells that migrate per explant per unit time, thus hindering the ability to gather sufficient numbers of cells for comprehensive experimentation. For this reason, we have modified an extracellular matrix digestion technique in current use for the isolation of microvascular endothelial cells to isolate human trabecular meshwork cells. This procedure is both efficient and rapid for isolating large numbers of trabecular meshwork cells and results in the availability of trabecular meshwork cells in sufficient quantities for subsequent experimentation. PMID:7587308

  2. Single Particle Tracking Reveals Biphasic Transport During Nanorod Magnetophoresis Through Extracellular Matrix

    PubMed Central

    Mair, L.O.; Superfine, R.

    2014-01-01

    Magnetic drug targeting has been proposed as a means of efficiently targeting drugs to tumors. However, the extracellular matrix (ECM) remains a significant barrier to long-range magnetophoretic transport through the tumor volume. While ensemble measurements of nanoparticle magnetophoresis have been reported, a single particle level understanding of magnetophoretic transport remains at large. We quantify nanorod magnetophoresis through ECM based on single particle observations. We find that smaller diameter particles achieve larger velocities through ECM despite experiencing smaller magnetic forces. Additionally, two interesting dynamics are elucidated. First, 18 nm diameter nanorods experience bimodal stick-slip motion through ECM during static field magnetophoresis, while similar bimodal transport is not observed for 55 nm nor 200 nm diameter nanorods. Second, smaller particles experience larger deviations in their orientation angle with respect to the magnetic field. This work elucidates important dynamics of nanoparticle transport through complex, porous biomaterials that may go unnoticed during ensemble measurements. PMID:24744160

  3. Cell-mediated fiber recruitment drives extracellular matrix mechanosensing in engineered fibrillar microenvironments

    PubMed Central

    Baker, Brendon M.; Trappmann, Britta; Wang, William Y.; Sakar, Mahmut S.; Kim, Iris L.; Shenoy, Vivek B.; Burdick, Jason A.; Chen, Christopher S.

    2015-01-01

    To investigate how cells sense stiffness in settings structurally similar to native extracellular matrices (ECM), we designed a synthetic fibrous material with tunable mechanics and user-defined architecture. In contrast to flat hydrogel surfaces, these fibrous materials recapitulated cell-matrix interactions observed with collagen matrices including stellate cell morphologies, cell-mediated realignment of fibers, and bulk contraction of the material. While increasing the stiffness of flat hydrogel surfaces induced mesenchymal stem cell spreading and proliferation, increasing fiber stiffness instead suppressed spreading and proliferation depending on network architecture. Lower fiber stiffness permitted active cellular forces to recruit nearby fibers, dynamically increasing ligand density at the cell surface and promoting the formation of focal adhesions and related signaling. These studies demonstrate a departure from the well-described relationship between material stiffness and spreading established with hydrogel surfaces, and introduce fiber recruitment as a novel mechanism by which cells probe and respond to mechanics in fibrillar matrices. PMID:26461445

  4. The dynamic architecture of photoreceptor ribbon synapses: Cytoskeletal, extracellular matrix, and intramembrane proteins

    PubMed Central

    MERCER, AARON J.; THORESON, WALLACE B.

    2012-01-01

    Rod and cone photoreceptors possess ribbon synapses that assist in the transmission of graded light responses to second-order bipolar and horizontal cells of the vertebrate retina. Proper functioning of the synapse requires the juxtaposition of presynaptic release sites immediately adjacent to postsynaptic receptors. In this review, we focus on the synaptic, cytoskeletal, and extracellular matrix proteins that help to organize photoreceptor ribbon synapses in the outer plexiform layer. We examine the proteins that foster the clustering of release proteins, calcium channels, and synaptic vesicles in the presynaptic terminals of photoreceptors adjacent to their postsynaptic contacts. Although many proteins interact with one another in the presynaptic terminal and synaptic cleft, these protein–protein interactions do not create a static and immutable structure. Instead, photoreceptor ribbon synapses are remarkably dynamic, exhibiting structural changes on both rapid and slow time scales. PMID:22192503

  5. Adhesion properties of Lactobacillus rhamnosus mucus-binding factor to mucin and extracellular matrix proteins.

    PubMed

    Nishiyama, Keita; Nakamata, Koichi; Ueno, Shintaro; Terao, Akari; Aryantini, Ni Putu Desy; Sujaya, I Nengah; Fukuda, Kenji; Urashima, Tadasu; Yamamoto, Yuji; Mukai, Takao

    2015-01-01

    We previously described potential probiotic Lactobacillus rhamnosus strains, isolated from fermented mare milk produced in Sumbawa Island, Indonesia, which showed high adhesion to porcine colonic mucin (PCM) and extracellular matrix (ECM) proteins. Recently, mucus-binding factor (MBF) was found in the GG strain of L. rhamnosus as a mucin-binding protein. In this study, we assessed the ability of recombinant MBF protein from the FSMM22 strain, one of the isolates of L. rhamnosus from fermented Sumbawa mare milk, to adhere to PCM and ECM proteins by overlay dot blot and Biacore assays. MBF bound to PCM, laminin, collagen IV, and fibronectin with submicromolar dissociation constants. Adhesion of the FSMM22 mbf mutant strain to PCM and ECM proteins was significantly less than that of the wild-type strain. Collectively, these results suggested that MBF contribute to L. rhamnosus host colonization via mucin and ECM protein binding. PMID:25351253

  6. Diabetes-Induced Alterations in the Extracellular Matrix and Their Impact on Myocardial Function

    PubMed Central

    Law, Brittany; Fowlkes, Vennece; Goldsmith, Jack G.; Carver, Wayne; Goldsmith, Edie C.

    2014-01-01

    Diabetes is an increasing public health problem that is expected to escalate in the future due to the growing incidence of obesity in the western world.While this disease is well known for its devastating effects on the kidneys and vascular system, diabetic individuals can develop cardiac dysfunction, termed diabetic cardiomyopathy, in the absence of other cardiovascular risk factors such as hypertension or atherosclerosis. While much effort has gone into understanding the effects of elevated glucose or altered insulin sensitivity on cellular components within the heart, significant changes in the cardiac extracellular matrix (ECM) have also been noted. In this review article we highlight what is currently known regarding the effects diabetes has on both the expression and chemical modification of proteins within the ECM and how the fibrotic response often observed as a consequence of this disease can contribute to reduced cardiac function. PMID:22221857

  7. Bromelain Surface Modification Increases the Diffusion of Silica Nanoparticles in the Tumor Extracellular Matrix

    PubMed Central

    2015-01-01

    Tumor extracellular matrix (ECM) represents a major obstacle to the diffusion of therapeutics and drug delivery systems in cancer parenchyma. This biological barrier limits the efficacy of promising therapeutic approaches including the delivery of siRNA or agents intended for thermoablation. After extravasation due to the enhanced penetration and retention effect of tumor vasculature, typical nanotherapeutics are unable to reach the nonvascularized and anoxic regions deep within cancer parenchyma. Here, we developed a simple method to provide mesoporous silica nanoparticles (MSN) with a proteolytic surface. To this extent, we chose to conjugate MSN to Bromelain (Br–MSN), a crude enzymatic complex, purified from pineapple stems, that belongs to the peptidase papain family. This surface modification increased particle uptake in endothelial, macrophage, and cancer cell lines with minimal impact on cellular viability. Most importantly Br–MSN showed an increased ability to digest and diffuse in tumor ECM in vitro and in vivo. PMID:25119793

  8. The extracellular matrix of Volvox carteri: molecular structure of the cellular compartment.

    PubMed

    Ertl, H; Mengele, R; Wenzl, S; Engel, J; Sumper, M

    1989-12-01

    The extracellular matrix (ECM) of Volvox contains insoluble fibrous layers that surround individual cells at a distance to form contiguous cellular compartments. Using immunological techniques, we identified a sulfated surface glycoprotein (SSG 185) as the monomeric precursor of this substructure within the ECM. The primary structure of the SSG 185 poly-peptide chain has been derived from cDNA and genomic DNA. A central domain of the protein, 80 amino acid residues long, consists almost exclusively of hydroxyproline residues. The chemical structure of the highly sulfated polysaccharide covalently attached to SSG 185 has been determined by permethylation analysis. As revealed by EM, SSG 185 is a rod-shaped molecule with a 21-nm-long polysaccharide strand protruding from its central region. The chemical nature of the cross-links between SSG 185 monomers is discussed. PMID:2689458

  9. Electrospun polyvinyl alcohol-collagen-hydroxyapatite nanofibers: a biomimetic extracellular matrix for osteoblastic cells

    NASA Astrophysics Data System (ADS)

    Song, Wei; Markel, David C.; Wang, Sunxi; Shi, Tong; Mao, Guangzhao; Ren, Weiping

    2012-03-01

    The failure of prosthesis after total joint replacement is due to the lack of early implant osseointegration. In this study polyvinyl alcohol-collagen-hydroxyapatite (PVA-Col-HA) electrospun nanofibrous meshes were fabricated as a biomimetic bone-like extracellular matrix for the modification of orthopedic prosthetic surfaces. In order to reinforce the PVA nanofibers, HA nanorods and Type I collagen were incorporated into the nanofibers. We investigated the morphology, biodegradability, mechanical properties and biocompatibility of the prepared nanofibers. Our results showed these inorganic-organic blended nanofibers to be degradable in vitro. The encapsulated nano-HA and collagen interacted with the PVA content, reinforcing the hydrolytic resistance and mechanical properties of nanofibers that provided longer lasting stability. The encapsulated nano-HA and collagen also enhanced the adhesion and proliferation of murine bone cells (MC3T3) in vitro. We propose the PVA-Col-HA nanofibers might be promising modifying materials on implant surfaces for orthopedic applications.

  10. In situ cross-linked polysaccharide hydrogel as extracellular matrix mimics for antibiotics delivery.

    PubMed

    Zhao, Yu; Zhang, Xinge; Wang, Yanan; Wu, Zhongming; An, Jinxia; Lu, Zhentan; Mei, Lin; Li, Chaoxing

    2014-05-25

    Many synthetic hydrogels for drug delivery have been based on polyethylene glycol which is non-natural, non-biodegradable and only terminal-functionalizable. The polysaccharides dextran and chitosan not only are highly hydrophilic, biodegradable and pendant-functionalizable, but also more closely mimic the nature extracellular matrix glycosaminoglycans. Here, a biomimetic hydrogel based on chitosan and dextran was synthesized by the Michael addition reaction. The hydrogels have good swelling and cytocompatibility against NIH3T3. Moreover, vancomycin-loaded hydrogels were formed in situ, and could kill both Gram-positive bacteria and Gram-negative bacteria, indicating that the hydrogel as a wound dressing could provide protection against bacterial infection. Notably, the drug release was controlled via modifying the compositions. Therefore, the biomimetic polysaccharide hydrogels as a promising carrier have potential application for wound healing. PMID:24708953

  11. Role of Extracellular Matrix Signaling Cues in Modulating Cell Fate Commitment for Cardiovascular Tissue Engineering

    PubMed Central

    Nakayama, Karina H.; Hou, Luqia; Huang, Ngan F.

    2014-01-01

    It is generally agreed that engineered cardiovascular tissues require cellular interactions with the local milieu. Within the microenvironment, the extracellular matrix (ECM) is an important support structure that provides dynamic signaling cues in part through its chemical, physical, and mechanical properties. In response to ECM factors, cells activate biochemical and mechanotransduction pathways that modulate their survival, growth, migration, differentiation, and function. This review describes the role of ECM chemical composition, spatial patterning, and mechanical stimulation in the specification of cardiovascular lineages, with a focus on stem cell differentiation, direct transdifferentiation, and endothelial-to-mesenchymal transition. The translational application of ECMs will be discussed in the context of cardiovascular tissue engineering and regenerative medicine. PMID:24443420

  12. Engineering approaches for investigating tumor angiogenesis: exploiting the role of the extracellular matrix

    PubMed Central

    Hielscher, Abigail C.; Gerecht, Sharon

    2012-01-01

    A major paradigm shift in cancer research is the emergence of multidisciplinary approaches to investigate complex cell behaviors to elucidate the regulatory mechanisms and to identify therapeutic targets. Recently, efforts are focused on the engineering of complex in-vitro models, which more accurately recapitulate the growth and progression of cancer. These strategies have proven vital for investigating and targeting the events that control tumor angiogenesis. In this review, we explore how the emerging engineering approaches are being utilized to unlock the complex mechanisms regulating tumor angiogenesis. Emphasis is placed on models employing natural and synthetic biomaterials to generate scaffolds mimicking the extracellular matrix (ECM) which plays a critical role in angiogenesis. While the models presented in this review are revolutionary, improvements are still necessary and concepts for advancing and perfecting engineering approaches for modeling tumor angiogenesis are proposed. Overall, the marriage between disparate scientific fields is expected to yield significant improvements of our understanding and treatment of cancer. PMID:23172313

  13. Extracellular matrix-inspired growth factor delivery systems for bone regeneration

    SciTech Connect

    Martino, Mikaël M.; Briquez, Priscilla S.; Maruyama, Kenta; Hubbell, Jeffrey A.

    2015-04-17

    Growth factors are very promising molecules to enhance bone regeneration. However, their translation to clinical use has been seriously limited, facing issues related to safety and cost-effectiveness. These problems derive from the vastly supra-physiological doses of growth factor used without optimized delivery systems. Therefore, these issues have motivated the development of new delivery systems allowing better control of the spatio-temporal release and signaling of growth factors. Because the extracellular matrix (ECM) naturally plays a fundamental role in coordinating growth factor activity in vivo, a number of novel delivery systems have been inspired by the growth factor regulatory function of the ECM. After introducing the role of growth factors during the bone regeneration process, this review exposes different issues that growth factor-based therapies have encountered in the clinic and highlights recent delivery approaches based on the natural interaction between growth factor and the ECM.

  14. The extracellular matrix protein Del1 induces apoptosis via its epidermal growth factor motif.

    PubMed

    Kitano, Hisataka; Kokubun, Shinichiro; Hidai, Chiaki

    2010-03-19

    Mouse Del1 is an extracellular matrix protein mainly expressed in the developing embryo. Del1 has three EGF motifs and two discoidin domains. The second EGF motif reportedly contains an RGD sequence that binds to integrin receptors. Here, we provide evidence that Del1 protein induces cell death in vitro. Chromatin condensation and DNA laddering were observed, suggestive of apoptosis. The results of analysis using the TUNEL method and annexin V staining were also consistent with apoptosis. The apoptosis-inducing activity of Del1 could be mapped to the third EGF motif, which fitted the consensus sequence CX(D/N)XXXX(F/Y)XCXC, wherein the aspartic acid residue (D) could be beta-hydroxylated. As little as twenty-five picomolar of recombinant E3 could induce apoptosis. PMID:20171188

  15. Biotensegrity of the Extracellular Matrix: Physiology, Dynamic Mechanical Balance, and Implications in Oncology and Mechanotherapy

    PubMed Central

    Tadeo, Irene; Berbegall, Ana P.; Escudero, Luis M.; Álvaro, Tomás; Noguera, Rosa

    2014-01-01

    Cells have the capacity to convert mechanical stimuli into chemical changes. This process is based on the tensegrity principle, a mechanism of tensional integrity. To date, this principle has been demonstrated to act in physiological processes such as mechanotransduction and mechanosensing at different scales (from cell sensing through integrins to molecular mechanical interventions or even localized massage). The process involves intra- and extracellular components, including the participation of extracellular matrix (ECM) and microtubules that act as compression structures, and actin filaments which act as tension structures. The nucleus itself has its own tensegrity system which is implicated in cell proliferation, differentiation, and apoptosis. Despite present advances, only the tip of the iceberg has so far been uncovered regarding the role of ECM compounds in influencing biotensegrity in pathological processes. Groups of cells, together with the surrounding ground substance, are subject to different and specific forces that certainly influence biological processes. In this paper, we review the current knowledge on the role of ECM elements in determining biotensegrity in malignant processes and describe their implication in therapeutic response, resistance to chemo- and radiotherapy, and subsequent tumor progression. Original data based on the study of neuroblastic tumors will be provided. PMID:24624363

  16. Extracellular Matrix-Regulated Gene Expression RequiresCooperation of SWI/SNF and Transcription Factors

    SciTech Connect

    Xu, Ren; Spencer, Virginia A.; Bissell, Mina J.

    2006-05-25

    Extracellular cues play crucial roles in the transcriptional regulation of tissue-specific genes, but whether and how these signals lead to chromatin remodeling is not understood and subject to debate. Using chromatin immunoprecipitation (ChIP) assays and mammary-specific genes as models, we show here that extracellular matrix (ECM) molecules and prolactin cooperate to induce histone acetylation and binding of transcription factors and the SWI/SNF complex to the {beta}- and ?-casein promoters. Introduction of a dominant negative Brg1, an ATPase subunit of SWI/SNF complex, significantly reduced both {beta}- and ?-casein expression, suggesting that SWI/SNF-dependent chromatin remodeling is required for transcription of mammary-specific genes. ChIP analyses demonstrated that the ATPase activity of SWI/SNF is necessary for recruitment of RNA transcriptional machinery, but not for binding of transcription factors or for histone acetylation. Coimmunoprecipitation analyses showed that the SWI/SNF complex is associated with STAT5, C/EBP{beta}, and glucocorticoid receptor (GR). Thus, ECM- and prolactin-regulated transcription of the mammary-specific casein genes requires the concerted action of chromatin remodeling enzymes and transcription factors.

  17. Biotensegrity of the extracellular matrix: physiology, dynamic mechanical balance, and implications in oncology and mechanotherapy.

    PubMed

    Tadeo, Irene; Berbegall, Ana P; Escudero, Luis M; Alvaro, Tomás; Noguera, Rosa

    2014-01-01

    Cells have the capacity to convert mechanical stimuli into chemical changes. This process is based on the tensegrity principle, a mechanism of tensional integrity. To date, this principle has been demonstrated to act in physiological processes such as mechanotransduction and mechanosensing at different scales (from cell sensing through integrins to molecular mechanical interventions or even localized massage). The process involves intra- and extracellular components, including the participation of extracellular matrix (ECM) and microtubules that act as compression structures, and actin filaments which act as tension structures. The nucleus itself has its own tensegrity system which is implicated in cell proliferation, differentiation, and apoptosis. Despite present advances, only the tip of the iceberg has so far been uncovered regarding the role of ECM compounds in influencing biotensegrity in pathological processes. Groups of cells, together with the surrounding ground substance, are subject to different and specific forces that certainly influence biological processes. In this paper, we review the current knowledge on the role of ECM elements in determining biotensegrity in malignant processes and describe their implication in therapeutic response, resistance to chemo- and radiotherapy, and subsequent tumor progression. Original data based on the study of neuroblastic tumors will be provided. PMID:24624363

  18. Differential expression of extracellular matrix and growth factors by embryoid bodies in hydrodynamic and static cultures.

    PubMed

    Fridley, Krista M; Nair, Rekha; McDevitt, Todd C

    2014-12-01

    During development, cell fate specification and tissue development are orchestrated by the sequential presentation of soluble growth factors (GF) and extracellular matrix (ECM) molecules. Similarly, differentiation of stem cells in vitro relies upon the temporal presence of extracellular cues within the microenvironment. Hydrodynamic culture systems are not limited by volume restrictions and therefore offer several practical advantages for scalability over static cultures; however, hydrodynamic cultures expose cells to physical parameters not present in static culture, such as fluid shear stress and mass transfer through convective forces. In this study, the differences between static and hydrodynamic culture conditions on the expression of ECM and GF molecules during the differentiation of mouse embryonic stem cells were examined at both the gene and protein level. The expression of ECM and GF genes exhibited an early decrease in static cultures based on heat map and hierarchical clustering analysis and a relative delayed increase in hydrodynamic cultures. Although the temporal patterns of specific ECM and GF protein expression were comparable between static and hydrodynamic cultures, several notable differences in the magnitudes of expression were observed at similar time points. These results describe the establishment of an analytical framework that can be used to examine the expression patterns of ECM and GF molecules expressed by pluripotent stem cells undergoing differentiation as 3D multicellular aggregates under different culture conditions, and suggest that physical parameters of stem cell microenvironments can alter endogenous ECM and GF expression profiles that may, in turn, influence cell fate decisions. PMID:25423310

  19. Granzyme B degrades extracellular matrix and contributes to delayed wound closure in apolipoprotein E knockout mice

    PubMed Central

    Hiebert, P R; Wu, D; Granville, D J

    2013-01-01

    Chronic inflammation and excessive protease activity have a major role in the persistence of non-healing wounds. Granzyme B (GzmB) is a serine protease expressed during chronic inflammation that, in conjunction with perforin, has a well-established role in initiating apoptotic cell death. GzmB is also capable of acting extracellularly, independent of perforin and can degrade several extracellular matrix (ECM) proteins that are critical during wound healing. We used apolipoprotein E (ApoE) knockout (AKO) mice as a novel model of chronic inflammation and impaired wound healing to investigate the role of GzmB in chronic wounds. Wild-type and AKO mice were grown to 7 weeks (young) or 37 weeks (old) of age on a regular chow or high-fat diet (HFD), given a 1-cm diameter full thickness wound on their mid dorsum and allowed to heal for 16 days. Old AKO mice fed a HFD exhibited reduced wound closure, delayed contraction, chronic inflammation and altered ECM remodeling. Conversely, GzmB/ApoE double knockout mice displayed improved wound closure and contraction rates. In addition, murine GzmB was found to degrade both fibronectin and vitronectin derived from healthy mouse granulation tissue. In addition, GzmB-mediated degradation of fibronectin generated a fragment similar in size to that observed in non-healing mouse wounds. These results provide the first direct evidence that GzmB contributes to chronic wound healing in part through degradation of ECM. PMID:23912712

  20. Extracellular matrix production and calcium carbonate precipitation by coral cells in vitro.

    PubMed

    Helman, Yael; Natale, Frank; Sherrell, Robert M; Lavigne, Michèle; Starovoytov, Valentin; Gorbunov, Maxim Y; Falkowski, Paul G

    2008-01-01

    The evolution of multicellularity in animals required the production of extracellular matrices that serve to spatially organize cells according to function. In corals, three matrices are involved in spatial organization: (i) an organic ECM, which facilitates cell-cell and cell-substrate adhesion; (ii) a skeletal organic matrix (SOM), which facilitates controlled deposition of a calcium carbonate skeleton; and (iii) the calcium carbonate skeleton itself, which provides the structural support for the 3D organization of coral colonies. In this report, we examine the production of these three matrices by using an in vitro culturing system for coral cells. In this system, which significantly facilitates studies of coral cell physiology, we demonstrate in vitro excretion of ECM by primary (nondividing) tissue cultures of both soft (Xenia elongata) and hard (Montipora digitata) corals. There are structural differences between the ECM produced by X. elongata cell cultures and that of M. digitata, and ascorbic acid, a critical cofactor for proline hydroxylation, significantly increased the production of collagen in the ECM of the latter species. We further demonstrate in vitro production of SOM and extracellular mineralized particles in cell cultures of M. digitata. Inductively coupled plasma mass spectrometry analysis of Sr/Ca ratios revealed the particles to be aragonite. De novo calcification was confirmed by following the incorporation of (45)Ca into acid labile macromolecules. Our results demonstrate the ability of isolated, differentiated coral cells to undergo fundamental processes required for multicellular organization. PMID:18162537

  1. Overview of the Matrisome—An Inventory of Extracellular Matrix Constituents and Functions

    PubMed Central

    Hynes, Richard O.; Naba, Alexandra

    2012-01-01

    Completion of genome sequences for many organisms allows a reasonably complete definition of the complement of extracellular matrix (ECM) proteins. In mammals this “core matrisome” comprises ∼300 proteins. In addition there are large numbers of ECM-modifying enzymes, ECM-binding growth factors, and other ECM-associated proteins. These different categories of ECM and ECM-associated proteins cooperate to assemble and remodel extracellular matrices and bind to cells through ECM receptors. Together with receptors for ECM-bound growth factors, they provide multiple inputs into cells to control survival, proliferation, differentiation, shape, polarity, and motility of cells. The evolution of ECM proteins was key in the transition to multicellularity, the arrangement of cells into tissue layers, and the elaboration of novel structures during vertebrate evolution. This key role of ECM is reflected in the diversity of ECM proteins and the modular domain structures of ECM proteins both allow their multiple interactions and, during evolution, development of novel protein architectures. PMID:21937732

  2. Role of extracellular matrix proteins and their receptors in the development of the vertebrate neuromuscular junction.

    PubMed

    Singhal, Neha; Martin, Paul T

    2011-11-01

    The vertebrate neuromuscular junction (NMJ) remains the best-studied model for understanding the mechanisms involved in synaptogenesis, due to its relatively large size, its simplicity of patterning, and its unparalleled experimental accessibility. During neuromuscular development, each skeletal myofiber secretes and deposits around its extracellular surface an assemblage of extracellular matrix (ECM) proteins that ultimately form a basal lamina. This is also the case at the NMJ, where the motor nerve contributes additional factors. Before most of the current molecular components were known, it was clear that the synaptic ECM of adult skeletal muscles was unique in composition and contained factors sufficient to induce the differentiation of both pre- and postsynaptic membranes. Biochemical, genetic, and microscopy studies have confirmed that agrin, laminin (221, 421, and 521), collagen IV (α3-α6), collagen XIII, perlecan, and the ColQ-bound form of acetylcholinesterase are all synaptic ECM proteins with important roles in neuromuscular development. The roles of their many potential receptors and/or binding proteins have been more difficult to assess at the genetic level due to the complexity of membrane interactions with these large proteins, but roles for MuSK-LRP4 in agrin signaling and for integrins, dystroglycan, and voltage-gated calcium channels in laminin-dependent phenotypes have been identified. Synaptic ECM proteins and their receptors are involved in almost all aspects of synaptic development, including synaptic initiation, topography, ultrastructure, maturation, stability, and transmission. PMID:21766463

  3. The widely expressed extracellular matrix protein SMOC-2 promotes keratinocyte attachment and migration

    SciTech Connect

    Maier, Silke; Paulsson, Mats; Hartmann, Ursula

    2008-08-01

    SMOC-2 is a recently discovered member of the BM-40/SPARC/osteonectin family of extracellular multidomain proteins of so far unknown function. While we have shown earlier that the homologous protein SMOC-1 is associated with basement membranes, in this study we demonstrate that, in the mouse, SMOC-2 could be detected in a large number of non-basement membrane localizations, often showing a diffuse tissue distribution. A more distinct expression pattern was seen in skin where SMOC-2 is mainly present in the basal layers of the epidermis. Functionally, recombinant SMOC-2 stimulated attachment of primary epidermal cells as well as several epidermal-derived cell lines but had no effect on the attachment of non-epidermal cells. Inhibition experiments using blocking antibodies against individual integrin subunits allowed the identification of {alpha}v{beta}6 and {alpha}v{beta}1 integrins as important cellular receptors for SMOC-2. Cell attachment as well as the formation of focal adhesions could be attributed to the extracellular calcium-binding domain. The calcium-binding domain also stimulated migration, but not proliferation of keratinocyte-like HaCaT cells. We conclude that SMOC-2, like other members of the BM40/SPARC family, acts as a regulator of cell-matrix interactions.

  4. Extracellular compartments in matrix morphogenesis: collagen fibril, bundle, and lamellar formation by corneal fibroblasts.

    PubMed

    Birk, D E; Trelstad, R L

    1984-12-01

    The regulation of collagen fibril, bundle, and lamella formation by the corneal fibroblasts, as well as the organization of these elements into an orthogonal stroma, was studied by transmission electron microscopy and high voltage electron microscopy. Transmission and high voltage electron microscopy of chick embryo corneas each demonstrated a series of unique extracellular compartments. Collagen fibrillogenesis occurred within small surface recesses. These small recesses usually contained between 5 and 12 collagen fibrils with typically mature diameters and constant intrafibrillar spacing. The lateral fusion of the recesses resulted in larger recesses and consequent formation of prominent cell surface foldings. Within these surface foldings, bundles that contained 50-100 collagen fibrils were formed. The surface foldings continued to fuse and the cell surface retracted, forming large surface-associated compartments in which bundles coalesced to form lamellae. High voltage electron microscopy of 0.5 micron sections cut parallel to the corneal surface revealed that the corneal fibroblasts and their processes had two major axes at approximately right angles to one another. The surface compartments involved in the production of the corneal stroma were aligned along the fibroblast axes and the orthogonality of the cell was in register with that of the extracellular matrix. In this manner, corneal fibroblasts formed collagen fibrils, bundles, and lamellae within a controlled environment and thereby determined the architecture of the corneal stroma by the configuration of the cell and its associated compartments. PMID:6542105

  5. Biological conduits combining bone marrow mesenchymal stem cells and extracellular matrix to treat long-segment sciatic nerve defects

    PubMed Central

    Wang, Yang; Li, Zheng-wei; Luo, Min; Li, Ya-jun; Zhang, Ke-qiang

    2015-01-01

    The transplantation of polylactic glycolic acid conduits combining bone marrow mesenchymal stem cells and extracellular matrix gel for the repair of sciatic nerve injury is effective in some respects, but few data comparing the biomechanical factors related to the sciatic nerve are available. In the present study, rabbit models of 10-mm sciatic nerve defects were prepared. The rabbit models were repaired with autologous nerve, a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells, or a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel. After 24 weeks, mechanical testing was performed to determine the stress relaxation and creep parameters. Following sciatic nerve injury, the magnitudes of the stress decrease and strain increase at 7,200 seconds were largest in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group, followed by the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group, and then the autologous nerve group. Hematoxylin-eosin staining demonstrated that compared with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group and the autologous nerve group, a more complete sciatic nerve regeneration was found, including good myelination, regularly arranged nerve fibers, and a completely degraded and resorbed conduit, in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group. These results indicate that bridging 10-mm sciatic nerve defects with a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel construct increases the stress relaxation under a constant strain, reducing anastomotic tension. Large elongations under a constant physiological load can limit the anastomotic opening and shift, which is beneficial for the regeneration and functional reconstruction of sciatic nerve. Better regeneration was

  6. Ultrasound Technologies for the Spatial Patterning of Cells and Extracellular Matrix Proteins and the Vascularization of Engineered Tissue

    NASA Astrophysics Data System (ADS)

    Garvin, Kelley A.

    Technological advancements in the field of tissue engineering could save the lives of thousands of organ transplant patients who die each year while waiting for donor organs. Currently, two of the primary challenges preventing tissue engineers from developing functional replacement tissues and organs are the need to recreate complex cell and extracellular microenvironments and to vascularize the tissue to maintain cell viability and function. Ultrasound is a form of mechanical energy that can noninvasively and nondestructively interact with tissues at the cell and protein level. In this thesis, novel ultrasound-based technologies were developed for the spatial patterning of cells and extracellular matrix proteins and the vascularization of three-dimensional engineered tissue constructs. Acoustic radiation forces associated with ultrasound standing wave fields were utilized to noninvasively control the spatial organization of cells and cell-bound extracellular matrix proteins within collagen-based engineered tissue. Additionally, ultrasound induced thermal mechanisms were exploited to site-specifically pattern various extracellular matrix collagen microstructures within a single engineered tissue construct. Finally, ultrasound standing wave field technology was used to promote the rapid and extensive vascularization of three-dimensional tissue constructs. As such, the ultrasound technologies developed in these studies have the potential to provide the field of tissue engineering with novel strategies to spatially pattern cells and extracellular matrix components and to vascularize engineered tissue, and thus, could advance the fabrication of functional replacement tissues and organs in the field of tissue engineering.

  7. Gene Expression in Human Hippocampus from Cocaine Abusers Identifies Genes which Regulate Extracellular Matrix Remodeling

    PubMed Central

    Mash, Deborah C.; ffrench-Mullen, Jarlath; Adi, Nikhil; Qin, Yujing; Buck, Andrew; Pablo, John

    2007-01-01

    The chronic effects of cocaine abuse on brain structure and function are blamed for the inability of most addicts to remain abstinent. Part of the difficulty in preventing relapse is the persisting memory of the intense euphoria or cocaine “rush”. Most abused drugs and alcohol induce neuroplastic changes in brain pathways subserving emotion and cognition. Such changes may account for the consolidation and structural reconfiguration of synaptic connections with exposure to cocaine. Adaptive hippocampal plasticity could be related to specific patterns of gene expression with chronic cocaine abuse. Here, we compare gene expression profiles in the human hippocampus from cocaine addicts and age-matched drug-free control subjects. Cocaine abusers had 151 gene transcripts upregulated, while 91 gene transcripts were downregulated. Topping the list of cocaine-regulated transcripts was RECK in the human hippocampus (FC = 2.0; p<0.05). RECK is a membrane-anchored MMP inhibitor that is implicated in the coordinated regulation of extracellular matrix integrity and angiogenesis. In keeping with elevated RECK expression, active MMP9 protein levels were decreased in the hippocampus from cocaine abusers. Pathway analysis identified other genes regulated by cocaine that code for proteins involved in the remodeling of the cytomatrix and synaptic connections and the inhibition of blood vessel proliferation (PCDH8, LAMB1, ITGB6, CTGF and EphB4). The observed microarray phenotype in the human hippocampus identified RECK and other region-specific genes that may promote long-lasting structural changes with repeated cocaine abuse. Extracellular matrix remodeling in the hippocampus may be a persisting effect of chronic abuse that contributes to the compulsive and relapsing nature of cocaine addiction. PMID:18000554

  8. Association of cell surface heparan sulfate proteoglycans of Schwann cells with extracellular matrix proteins.

    PubMed

    Carey, D J; Crumbling, D M; Stahl, R C; Evans, D M

    1990-11-25

    The terminal differentiation of Schwann cells is dependent on contact with basement membrane. The present study was undertaken to investigate the role of cell surface heparan sulfate proteoglycans (HSPGs) in mediating Schwann cell responses to extracellular matrix contact. Phosphatidylinositol-specific phospholipase C-releasable cell surface HSPGs purified from cultures of neonatal rat Schwann cells were subjected to affinity chromatography on immobilized laminin and fibronectin. Binding of the HSPG to both affinity matrices was observed. The strength of the association, however, was sensitive to the ionic strength of the buffer. In 0.1 M Tris-HCl, HSPG binding was essentially irreversible whereas in physiological ionic strength buffer (e.g. 0.142 M NaCl, 10 mM Tris), weaker binding was detected as a delay in elution of the HSPG from the affinity columns. Further studies of HSPG-laminin binding suggested that the binding was mediated by the glycosaminoglycan chains of the proteoglycans. Results of equilibrium gel filtration chromatography provided additional evidence for a reversible association of the HSPG and laminin with a Kd of approximately 1 x 10(-6) M. When Schwann cells were plated on plastic dishes coated with laminin, the cells attached and extended long slender processes. Inclusion of heparin, but not chondroitin sulfate, in the assay medium resulted in partial inhibition of process extension, but at concentrations of heparin which were higher than that needed to disrupt laminin-HSPG association in vitro. Addition of anti-integrin receptor antibodies resulted in more extensive inhibition of laminin-dependent process extension. Anti-integrin antibodies plus heparin essentially totally inhibited laminin-dependent process extension. These results demonstrate that cell surface HSPGs are capable of reversible association with extracellular matrix molecules and suggest that HSPG-laminin interactions play a role in laminin-dependent Schwann cell spreading. PMID

  9. PTEN Regulates Renal Extracellular Matrix Deposit via Increased CTGF in Diabetes Mellitus.

    PubMed

    Zhu, Lin; Zhao, Song; Liu, Shuxia; Liu, Qingjuan; Li, Fan; Hao, Jun

    2016-05-01

    Extracellular matrix accumulation and fibrosis are the features of diabetic nephropathy. PI3K (phosphatidylinositol 3-kinase)/Akt (protein kinase B) signal pathway and its inhibitor PTEN (phosphatase and tensin homolog deleted on chromosome 10) are revealed to modulate renal fibrosis. However, the exact mechanism is still not well known. In the present study we found that compared with normal mice, diabetic mice showed decreased PTEN, increased phospho-Akt (Ser 473), phospho-Akt (Thr 308), CTGF (connective tissue growth factor), α-SMA (α-smooth muscle actin), and matricellular protein in kidney. Knocking down of PTEN caused an increase in phospho-Akt (Ser 473), phospho-Akt (Thr 308), CTGF, secreted fibronectin, and secreted Col 3 in HKC cells (human renal tubular epithelial cells). Again, in vitro experiment revealed 1.89, 2.18, 1.92, 3.06, 2.06-fold increases of phospho-Akt (Ser 473), phospho-Akt (Thr 308), CTGF, secreted fibronectin, and secreted Col 3 in high glucose-stimulated HKC cells in comparison with normal control cells. Furthermore, knocking down of CTGF reversed increased secreted fibronectin and Col 3 in high glucose-treated HKC cells. Moreover, transfection of PTEN expression vector prevented high glucose-caused these changes in HKC cells. Especially, CTGF expression, secretion of fibronectin and Col 3 were, respectively, decreased by 38.81, 53.85, and 39.12%. The treatment of LY294002 inhibited phospho-Akt (Ser 473) and phospho-Akt (Thr 308) expression followed by decreased CTGF, secretory fibronectin and secretory Col 3 in high glucose-treated HKC cells. In the end our study suggests that PTEN regulates renal extracellular matrix production via activated Akt and increased CTGF in diabetes mellitus. J. Cell. Biochem. 117: 1187-1198, 2016. © 2015 Wiley Periodicals, Inc. PMID:26447680

  10. Cell surface localization of heparanase on macrophages regulates degradation of extracellular matrix heparan sulfate.

    PubMed

    Sasaki, Norihiko; Higashi, Nobuaki; Taka, Tomohiro; Nakajima, Motowo; Irimura, Tatsuro

    2004-03-15

    Extravasation of peripheral blood monocytes through vascular basement membranes requires degradation of extracellular matrix components including heparan sulfate proteoglycans (HSPGs). Heparanase, the heparan sulfate-specific endo-beta-glucuronidase, has previously been shown to be a key enzyme in melanoma invasion, yet its involvement in monocyte extravasation has not been elucidated. We examined a potential regulatory mechanism of heparanase in HSPG degradation and transmigration through basement membranes in leukocyte trafficking using human promonocytic leukemia U937 and THP-1 cells. PMA-treated cells were shown to degrade 35S-sulfated HSPG in endothelial extracellular matrix into fragments of an approximate molecular mass of 5 kDa. This was not found with untreated cells. The gene expression levels of heparanase or the enzyme activity of the amount of cell lysates were no different between untreated and treated cells. Immunocytochemical staining with anti-heparanase mAb revealed pericellular distribution of heparanase in PMA-treated cells but not in untreated cells. Cell surface heparanase capped into a restricted area on PMA-treated cells when they were allowed to adhere. Addition of a chemoattractant fMLP induced polarization of the PMA-treated cells and heparanase redistribution at the leading edge of migration. Therefore a major regulatory process of heparanase activity in the cells seems to be surface expression and capping of the enzyme. Addition of the anti-heparanase Ab significantly inhibited enzymatic activity and transmigration of the PMA-treated cells, suggesting that the cell surface redistribution of heparanase is involved in monocyte extravasation through basement membranes. PMID:15004189

  11. Protein Modification by Deamidation Indicates Variations in Joint Extracellular Matrix Turnover*

    PubMed Central

    Catterall, Jonathan B.; Hsueh, Ming F.; Stabler, Thomas V.; McCudden, Christopher R.; Bolognesi, Michael; Zura, Robert; Jordan, Joanne M.; Renner, Jordan B.; Feng, Sheng; Kraus, Virginia B.

    2012-01-01

    As extracellular proteins age, they undergo and accumulate nonenzymatic post-translational modifications that cannot be repaired. We hypothesized that these could be used to systemically monitor loss of extracellular matrix due to chronic arthritic diseases such as osteoarthritis (OA). To test this, we predicted sites of deamidation in cartilage oligomeric matrix protein (COMP) and confirmed, by mass spectroscopy, the presence of deamidated (Asp64) and native (Asn64) COMP epitopes (mean 0.95% deamidated COMP (D-COMP) relative to native COMP) in cartilage. An Asp64, D-COMP-specific ELISA was developed using a newly created monoclonal antibody 6-1A12. In a joint replacement study, serum D-COMP (p = 0.017), but not total COMP (p = 0.5), declined significantly after replacement demonstrating a joint tissue source for D-COMP. In analyses of 450 participants from the Johnston County Osteoarthritis Project controlled for age, gender, and race, D-COMP was associated with radiographic hip (p < 0.0001) but not knee (p = 0.95) OA severity. In contrast, total COMP was associated with radiographic knee (p < 0.0001) but not hip (p = 0.47) OA severity. D-COMP was higher in soluble proteins extracted from hip cartilage proximal to OA lesions compared with remote from lesions (p = 0.007) or lesional and remote OA knee (p < 0.01) cartilage. Total COMP in cartilage did not vary by joint site or proximity to the lesion. This study demonstrates the presence of D-COMP in articular cartilage and the systemic circulation, and to our knowledge, it is the first biomarker to show specificity for a particular joint site. We believe that enrichment of deamidated epitope in hip OA cartilage indicates a lesser repair response of hip OA compared with knee OA cartilage. PMID:22179616

  12. The Role of the Extracellular Matrix Protein Mindin in Airway Response to Environmental Airways Injury

    PubMed Central

    Frush, Sarah; Li, Zhuowei; Potts, Erin N.; Du, Wanglei; Eu, Jerry P.; Garantziotis, Stavros; He, You-Wen; Foster, W. Michael

    2011-01-01

    Background: Our previous work demonstrated that the extracellular matrix protein mindin contributes to allergic airways disease. However, the role of mindin in nonallergic airways disease has not previously been explored. Objectives: We hypothesized that mindin would contribute to airways disease after inhalation of either lipopolysaccharide (LPS) or ozone. Methods: We exposed C57BL/6J and mindin-deficient (–/–) mice to aerosolized LPS (0.9 μg/m3 for 2.5 hr), saline, ozone (1 ppm for 3 hr), or filtered air (FA). All mice were evaluated 4 hr after LPS/saline 
exposure or 24 hr after ozone/FA exposure. We characterized the physiological and biological responses by analysis of airway hyperresponsiveness (AHR) with a computer-controlled small-animal ventilator (FlexiVent), inflammatory cellular recruitment, total protein in bronchoalveolar lavage fluid (BALF), proinflammatory cytokine profiling, and ex vivo bronchial ring studies. Results: After inhalation of LPS, mindin–/– mice demonstrated significantly reduced total cell and neutrophil recruitment into the airspace compared with their wild-type counterparts. Mindin–/– mice also exhibited reduced proinflammatory cytokine production and lower AHR to methacholine challenge by FlexiVent. After inhalation of ozone, mice had no detectible differences in cellular inflammation or total BALF protein dependent on mindin. However, mindin–/– mice were protected from increased proinflammatory cytokine production and AHR compared with their C57BL/6J counterparts. After ozone exposure, bronchial rings derived from mindin–/– mice demonstrated reduced constriction in response to carbachol. Conclusions: These data demonstrate that the extracellular matrix protein mindin modifies the airway response to both LPS and ozone. Our data support a conserved role of mindin in production of proinflammatory cytokines and the development of AHR in two divergent models of reactive airways disease, as well as a role of

  13. A polygenic burden of rare variants across extracellular matrix genes among individuals with adolescent idiopathic scoliosis.

    PubMed

    Haller, Gabe; Alvarado, David; Mccall, Kevin; Yang, Ping; Cruchaga, Carlos; Harms, Matthew; Goate, Alison; Willing, Marcia; Morcuende, Jose A; Baschal, Erin; Miller, Nancy H; Wise, Carol; Dobbs, Matthew B; Gurnett, Christina A

    2016-01-01

    Adolescent idiopathic scoliosis (AIS) is a complex inherited spinal deformity whose etiology has been elusive. While common genetic variants are associated with AIS, they explain only a small portion of disease risk. To explore the role of rare variants in AIS susceptibility, exome sequence data of 391 severe AIS cases and 843 controls of European ancestry were analyzed using a pathway burden analysis in which variants are first collapsed at the gene level then by Gene Ontology terms. Novel non-synonymous/splice-site variants in extracellular matrix genes were significantly enriched in AIS cases compared with controls (P = 6 × 10(-9), OR = 1.7, CI = 1.4-2.0). Specifically, novel variants in musculoskeletal collagen genes were present in 32% (126/391) of AIS cases compared with 17% (146/843) of in-house controls and 18% (780/4300) of EVS controls (P = 1 × 10(-9), OR = 1.9, CI = 1.6-2.4). Targeted resequencing of six collagen genes replicated this association in combined 919 AIS cases (P = 3 × 10(-12), OR = 2.2, CI = 1.8-2.7) and revealed a highly significant single-gene association with COL11A2 (P = 6 × 10(-9), OR = 3.8, CI = 2.6-7.2). Importantly, AIS cases harbor mainly non-glycine missense mutations and lack the clinical features of monogenic musculoskeletal collagenopathies. Overall, our study reveals a complex genetic architecture of AIS in which a polygenic burden of rare variants across extracellular matrix genes contributes strongly to risk. PMID:26566670

  14. Nitrotyrosine immunostaining correlates with increased extracellular matrix: evidence of postplacental hypoxia.

    PubMed

    Stanek, J; Eis, A L; Myatt, L

    2001-04-01

    Nitrotyrosine residues (NT), an index of oxidative stress arising from peroxynitrite formation and action, are found in placental vasculature of pregnancies complicated by pre-eclampsia (PE) or pregestational insulin-dependent diabetes mellitus (IDDM). This study correlates conventional placental pathology with NT immunostaining in 20 cases of perinatal mortality (13 stillbirths and seven cases of neonatal mortality) associated with PE, IDDM, amniotic fluid infection syndrome (AFIS), or from fetal/neonatal demise not related to these conditions (congenital anomalies) (n = five/group). Patients with PE have more decidual arteriolopathy and Tenney-Parker change, while patients with IDDM and ascending infection have more villous cytotrophoblastic hyperplasia. Archival paraffin-embedded placental sections were immunostained for NT for correlation with clinical features and H&E histological findings. The intensity of immunostaining for NT varied from absent (n = 7) to 1+ (n = 5) or 2+ (n = 8). All eight placentae with 2+ staining showed increased villous extracellular matrix (ECM), compared to none of five with 1+ staining and two of seven with no staining (chi2 = 14.3, P = 0.001). There was no statistically significant difference in the percentage of stem villi with luminal vascular abnormalities (5.7 vs 10 vs 35.7 per cent, F = 2.3, P = 0.1). Our data show that increased production of reactive oxygen species by placental tissue may be associated with increased extracellular matrix, itself produced by fibroblasts under the influence of oxygen. NT immunostaining may therefore help differentiate those cases of perinatal morbidity/mortality associated with post-placental hypoxia provided that the secondary impact of intrauterine fetal death can be excluded by future studies. PMID:11312630

  15. Co-transfection of decorin and interleukin-10 modulates pro-fibrotic extracellular matrix gene expression in human tenocyte culture

    PubMed Central

    Abbah, Sunny A.; Thomas, Dilip; Browne, Shane; O’Brien, Timothy; Pandit, Abhay; Zeugolis, Dimitrios I.

    2016-01-01

    Extracellular matrix synthesis and remodelling are driven by increased activity of transforming growth factor beta 1 (TGF-β1). In tendon tissue repair, increased activity of TGF-β1 leads to progressive fibrosis. Decorin (DCN) and interleukin 10 (IL-10) antagonise pathological collagen synthesis by exerting a neutralising effect via downregulation of TGF-β1. Herein, we report that the delivery of DCN and IL-10 transgenes from a collagen hydrogel system supresses the constitutive expression of TGF-β1 and a range of pro-fibrotic extracellular matrix genes. PMID:26860065

  16. Co-transfection of decorin and interleukin-10 modulates pro-fibrotic extracellular matrix gene expression in human tenocyte culture

    NASA Astrophysics Data System (ADS)

    Abbah, Sunny A.; Thomas, Dilip; Browne, Shane; O'Brien, Timothy; Pandit, Abhay; Zeugolis, Dimitrios I.

    2016-02-01

    Extracellular matrix synthesis and remodelling are driven by increased activity of transforming growth factor beta 1 (TGF-β1). In tendon tissue repair, increased activity of TGF-β1 leads to progressive fibrosis. Decorin (DCN) and interleukin 10 (IL-10) antagonise pathological collagen synthesis by exerting a neutralising effect via downregulation of TGF-β1. Herein, we report that the delivery of DCN and IL-10 transgenes from a collagen hydrogel system supresses the constitutive expression of TGF-β1 and a range of pro-fibrotic extracellular matrix genes.

  17. Expression of extracellular matrix proteins in cervical squamous cell carcinoma--a clinicopathological study.

    PubMed Central

    Goldberg, I; Davidson, B; Lerner-Geva, L; Gotlieb, W H; Ben-Baruch, G; Novikov, I; Kopolovic, J

    1998-01-01

    AIM: To evaluate the intracellular and peritumoral expression of matrix proteins in squamous cell carcinoma of the uterine cervix using immunohistochemistry. METHODS: 71 squamous cell carcinomas and 10 controls were stained for laminin, fibronectin, and collagen IV. Cytoplasmic staining in tumour cells and peritumoral deposition of matrix proteins were evaluated. The association between staining results and patient age, tumour stage, histological grade, and survival was studied. RESULTS: Positive cytoplasmic staining for laminin, fibronectin, and collagen IV was observed in 17 (23.9%), 27 (38%), and 10 (14.1%) cases, respectively. Staining for laminin was most pronounced in the invasive front of tumour islands, while for fibronectin and collagen IV it appeared to be diffuse. Peritumoral staining for laminin and collagen IV was detected in 12 cases (16.9%). Early stage (Ia1-Ia2) tumours were uniformly negative for all three proteins. Cytoplasmic staining for laminin correlated with positive staining for fibronectin and collagen IV, and with the presence of a peritumoral deposition of collagen IV and laminin. There was no correlation with any of the three markers between staining results and patient age, stage, grade, or survival. CONCLUSIONS: Expression of extracellular matrix proteins in some cervical squamous cell carcinomas might reflect the enhanced ability of these tumours to modify the peritumoral stroma. This ability seems to be absent in early stage tumours. The correlation between intracytoplasmic and peritumoral expression of matrix proteins supports the evidence of their synthesis by tumour cells. However, this property did not correlate with disease outcome in this study. Images PMID:10023343

  18. Lumican, an extracellular matrix proteoglycan, is a novel requisite for hepatic fibrosis

    PubMed Central

    Krishnan, Anuradha; Li, Xia; Kao, Winston Whei-Yang; Viker, Kimberly; Butters, Kim; Masuoka, Howard; Knudsen, Bruce; Gores, Gregory; Charlton, Michael

    2013-01-01

    Lumican, an extracellular matrix proteoglycan was previously shown to be upregulated with increasing severity of nonalcoholic steatohepatitis (NASH). Although lumican is involved in collagen fibrillogenesis in extra-hepatic tissues, little is known about the role of lumican in hepatic disease. We therefore determined lumican expression in etiologies other than clinical NASH. Our results indicated that lumican is upregulated in clinical samples of hepatitis C virus infection, in experimental rodent models of chronic and acute liver injury and could additionally be induced in vitro in response to the pro-fibrotic cytokine transforming growth factorβ1 (TGFβ 1) and to lipotoxic palmitic acid. Together, these results suggested a role for lumican in hepatic fibrosis. To investigate the functional role of lumican in hepatic fibrosis, lumican null (Null) and wild-type (WT) littermates were administered carbon tetrachloride intra-peritoneally. Serum and liver tissue were analyzed for indices of liver injury, fibrosis, matrix turnover, and proliferation. Hepatic fibrosis was greatly reduced in null animals (p<0.05). Paradoxically, gene expression of fibrosis-related genes such as TGFβ 1 and collagen 1 was numerically higher in null animals though statistically insignificant from WT animals. On the other hand, αsmooth muscle actin expression (α-SMA), a marker for activated fibroblasts, the main contributors of collagen production was significantly higher (p<0.05) in null animals as compared with WT littermates. Among the matrix metalloproteases (MMP), MMP13 was significantly increased (p<0.05) in null animals. Ultra-structural imaging indicated differences in the organization and spatial distribution of hepatic collagen fibrils of null and WT mice. Cell proliferation was significantly increased (p<0.05) in null animals. We conclude that lumican is a prerequisite for hepatic fibrosis. The protective effect of lumican deficiency in hepatic fibrosis appears to be downstream

  19. Expression of extracellular matrix metalloproteinase inducer and enhancement of the production of matrix metalloproteinase-1 in tongue squamous cell carcinoma.

    PubMed

    Cao, Z; Xiang, J; Li, C

    2009-08-01

    Recent studies have found that in addition to promoting cellular invasion, overexpression of metalloproteinase -1 (MMP-1) is associated with the initial stages of cancer development. Extracellular matrix metalloproteinase inducer (EMMPRIN), a transmembrane glycoprotein, has been reported to be highly expressed in tumor cells and induce production of MMPs from peritumor fibroblasts (PTFs) adjacent to the tumor cells. The expression of EMMPRIN in tongue squamous cell carcinoma (SCC) was investigated in this study. It was found that EMMPRIN was expressed at the cell membrane throughout the entire lesion in tongue SCC. Immunofluorescence staining localized EMMPRIN to the cell membrane in a highly invasive tongue SCC cell line (Tca 8113). EMMPRIN mRNA was expressed at a high level in Tca 8113, whereas MMP-1 mRNA was expressed in PTF but harder to be detected in Tca 8113. Co-culture of Tca 8113 with PTF stimulated production of MMP-1. EMMPRIN was highly expressed in tongue SCC, and could induce local production of MMP-1. These data indicate that EMMPRIN might play an important role in tongue SCC progression and invasion. PMID:19372030

  20. The regulation of growth and metabolism of kidney stem cell with regional specificity using extracellular matrix derived from kidney

    PubMed Central

    O’Neill, John D.; Freytes, Donald O.; Anandappa, Annabelle; Oliver, Juan A.; Vunjak-Novakovic, Gordana

    2013-01-01

    Native extracellular matrix (ECM) that is secreted and maintained by resident cells is of great interest for cell culture and cell delivery. We hypothesized that specialized bioengineered niches for stem cells can be established using ECM-derived scaffolding materials. Kidney was selected as a model system because of the high regional diversification of renal tissue matrix. By preparing the ECM from three specialized regions of the kidney (cortex, medulla, and papilla; whole kidney, heart, and bladder as controls) in three forms: (i) intact sheets of decellularized ECM, (ii) ECM hydrogels, and (iii) solubilized ECM, we investigated how the structure and composition of ECM affect the function of kidney stem cells (with mesenchymal stem cells, MSCs, as controls). All three forms of the ECM regulated KSC function, with differential structural and compositional effects. KSCs cultured on papilla ECM consistently displayed lower proliferation, higher metabolic activity, and differences in cell morphology, alignment, and structure formation as compared to KSCs on cortex and medulla ECM, effects not observed in corresponding MSC cultures. These data suggest that tissue- and region-specific ECM can provide an effective substrate for in vitro studies of therapeutic stem cells. PMID:24074840

  1. Maintenance of the Extracellular Matrix in Rat Anterior Pituitary Gland: Identification of Cells Expressing Tissue Inhibitors of Metalloproteinases

    PubMed Central

    Azuma, Morio; Tofrizal, Alimuddin; Maliza, Rita; Batchuluun, Khongorzul; Ramadhani, Dini; Syaidah, Rahimi; Tsukada, Takehiro; Fujiwara, Ken; Kikuchi, Motoshi; Horiguchi, Kotaro; Yashiro, Takashi

    2015-01-01

    The extracellular matrix (ECM) is important in creating cellular environments in tissues. Recent studies have demonstrated that ECM components are localized in anterior pituitary cells and affect cell activity. Thus, clarifying the mechanism responsible for ECM maintenance would improve understanding of gland function. Tissue inhibitors of metalloproteinases (TIMPs) are endogenous inhibitors of matrix metalloproteinases and participate in ECM degradation. In this study, we investigated whether cells expressing TIMPs are present in rat anterior pituitary gland. Reverse transcription polymerase chain reaction was used to analyze expression of the TIMP family (TIMP1-4), and cells producing TIMPs in the gland were identified by using in situ hybridization. Expression of TIMP1, TIMP2, and TIMP3 mRNAs was detected, and the TIMP-expressing cells were located in the gland. The TIMP-expressing cells were also investigated by means of double-staining with in situ hybridization and immunohistochemical techniques. Double-staining revealed that TIMP1 mRNA was expressed in folliculostellate cells. TIMP2 mRNA was detected in folliculostellate cells, prolactin cells, and thyroid-stimulating hormone cells. TIMP3 mRNA was identified in endothelial cells, pericytes, novel desmin-immunopositive perivascular cells, and folliculostellate cells. These findings indicate that TIMP1-, TIMP2-, and TIMP3-expressing cells are present in rat anterior pituitary gland and that they are involved in maintaining ECM components. PMID:26855451

  2. New methods to study the composition and structure of the extracellular matrix in natural and bioengineered tissues

    PubMed Central

    Schiller, Jürgen; Huster, Daniel

    2012-01-01

    The extracellular matrix (ECM) comprises a gel of numerous biopolymers that occurs in a multitude of biological tissues. The ECM provides the basic support and mechanical strength of skeletal tissue and is responsible for shape retention. At the same time, the ECM is responsible for the viscoelastic properties and the elasticity of soft tissues. As expected, there are several important diseases that affect and degenerate the ECM with severe consequences for its properties. Bioengineering is a promising approach to support the regenerative capacity of the body. Unfortunately, the biomechanical properties of bioengineered ECM often only poorly meet the standards of their native counterparts. Many bioengineered tissues are characterized by an increased glycosaminoglycan (GAG) but decreased collagen content. This leads to an enhanced water content that strongly alters the viscoelastic and thus the biomechanical properties. Therefore, compositional analysis is important to estimate the tissue quality. We will show that nuclear magnetic resonance (NMR) spectroscopy and soft-ionization mass spectrometry (MS) represent useful techniques for ECM research both in natural and bioengineered tissues. Both methods are strongly complimentary: while MS techniques such as matrix-assisted laser desorption and ionization (MALDI) are excellent and very sensitive analytical tools to determine the collagen and the GAG contents of tissues, NMR spectroscopy provides insight into the molecular architecture of the ECM, its dynamics and other important parameters such as the water content of the tissue as well as the diffusion of molecules within the ECM. PMID:23507863

  3. The regulation of growth and metabolism of kidney stem cells with regional specificity using extracellular matrix derived from kidney.

    PubMed

    O'Neill, John D; Freytes, Donald O; Anandappa, Annabelle J; Oliver, Juan A; Vunjak-Novakovic, Gordana V

    2013-12-01

    Native extracellular matrix (ECM) that is secreted and maintained by resident cells is of great interest for cell culture and cell delivery. We hypothesized that specialized bioengineered niches for stem cells can be established using ECM-derived scaffolding materials. Kidney was selected as a model system because of the high regional diversification of renal tissue matrix. By preparing the ECM from three specialized regions of the kidney (cortex, medulla, and papilla; whole kidney, heart, and bladder as controls) in three forms: (i) intact sheets of decellularized ECM, (ii) ECM hydrogels, and (iii) solubilized ECM, we investigated how the structure and composition of ECM affect the function of kidney stem cells (with mesenchymal stem cells, MSCs, as controls). All three forms of the ECM regulated KSC function, with differential structural and compositional effects. KSCs cultured on papilla ECM consistently displayed lower proliferation, higher metabolic activity, and differences in cell morphology, alignment, and structure formation as compared to KSCs on cortex and medulla ECM, effects not observed in corresponding MSC cultures. These data suggest that tissue- and region-specific ECM can provide an effective substrate for in vitro studies of therapeutic stem cells. PMID:24074840

  4. Physical, Spatial, and Molecular Aspects of Extracellular Matrix of In Vivo Niches and Artificial Scaffolds Relevant to Stem Cells Research.

    PubMed

    Akhmanova, Maria; Osidak, Egor; Domogatsky, Sergey; Rodin, Sergey; Domogatskaya, Anna

    2015-01-01

    Extracellular matrix can influence stem cell choices, such as self-renewal, quiescence, migration, proliferation, phenotype maintenance, differentiation, or apoptosis. Three aspects of extracellular matrix were extensively studied during the last decade: physical properties, spatial presentation of adhesive epitopes, and molecular complexity. Over 15 different parameters have been shown to influence stem cell choices. Physical aspects include stiffness (or elasticity), viscoelasticity, pore size, porosity, amplitude and frequency of static and dynamic deformations applied to the matrix. Spatial aspects include scaffold dimensionality (2D or 3D) and thickness; cell polarity; area, shape, and microscale topography of cell adhesion surface; epitope concentration, epitope clustering characteristics (number of epitopes per cluster, spacing between epitopes within cluster, spacing between separate clusters, cluster patterns, and level of disorder in epitope arrangement), and nanotopography. Biochemical characteristics of natural extracellular matrix molecules regard diversity and structural complexity of matrix molecules, affinity and specificity of epitope interaction with cell receptors, role of non-affinity domains, complexity of supramolecular organization, and co-signaling by growth factors or matrix epitopes. Synergy between several matrix aspects enables stem cells to retain their function in vivo and may be a key to generation of long-term, robust, and effective in vitro stem cell culture systems. PMID:26351461

  5. Physical, Spatial, and Molecular Aspects of Extracellular Matrix of In Vivo Niches and Artificial Scaffolds Relevant to Stem Cells Research

    PubMed Central

    Akhmanova, Maria; Osidak, Egor; Domogatsky, Sergey; Rodin, Sergey; Domogatskaya, Anna

    2015-01-01

    Extracellular matrix can influence stem cell choices, such as self-renewal, quiescence, migration, proliferation, phenotype maintenance, differentiation, or apoptosis. Three aspects of extracellular matrix were extensively studied during the last decade: physical properties, spatial presentation of adhesive epitopes, and molecular complexity. Over 15 different parameters have been shown to influence stem cell choices. Physical aspects include stiffness (or elasticity), viscoelasticity, pore size, porosity, amplitude and frequency of static and dynamic deformations applied to the matrix. Spatial aspects include scaffold dimensionality (2D or 3D) and thickness; cell polarity; area, shape, and microscale topography of cell adhesion surface; epitope concentration, epitope clustering characteristics (number of epitopes per cluster, spacing between epitopes within cluster, spacing between separate clusters, cluster patterns, and level of disorder in epitope arrangement), and nanotopography. Biochemical characteristics of natural extracellular matrix molecules regard diversity and structural complexity of matrix molecules, affinity and specificity of epitope interaction with cell receptors, role of non-affinity domains, complexity of supramolecular organization, and co-signaling by growth factors or matrix epitopes. Synergy between several matrix aspects enables stem cells to retain their function in vivo and may be a key to generation of long-term, robust, and effective in vitro stem cell culture systems. PMID:26351461

  6. A Collagen-based Scaffold Delivering Exogenous MicroRNA-29B to Modulate Extracellular Matrix Remodeling

    PubMed Central

    Monaghan, Michael; Browne, Shane; Schenke-Layland, Katja; Pandit, Abhay

    2014-01-01

    Directing appropriate extracellular matrix remodeling is a key aim of regenerative medicine strategies. Thus, antifibrotic interfering RNA (RNAi) therapy with exogenous microRNA (miR)-29B was proposed as a method to modulate extracellular matrix remodeling following cutaneous injury. It was hypothesized that delivery of miR-29B from a collagen scaffold will efficiently modulate the extracellular matrix remodeling response and reduce maladaptive remodeling such as aggressive deposition of collagen type I after injury. The release of RNA from the scaffold was assessed and its ability to silence collagen type I and collagen type III expression was evaluated in vitro. When primary fibroblasts were cultured with scaffolds doped with miR-29B, reduced levels of collagen type I and collagen type III mRNA expression were observed for up to 2 weeks of culture. When the scaffolds were applied to full thickness wounds in vivo, reduced wound contraction, improved collagen type III/I ratios and a significantly higher matrix metalloproteinase (MMP)-8: tissue inhibitor of metalloproteinase (TIMP)-1 ratio were detected when the scaffolds were functionalized with miR-29B. Furthermore, these effects were significantly influenced by the dose of miR-29B in the collagen scaffold (0.5 versus 5 μg). This study shows a potential of combining exogenous miRs with collagen scaffolds to improve extracellular matrix remodeling following injury. PMID:24402185

  7. Genetic Background is a Key Determinant of Glomerular Extracellular Matrix Composition and Organization.

    PubMed

    Randles, Michael J; Woolf, Adrian S; Huang, Jennifer L; Byron, Adam; Humphries, Jonathan D; Price, Karen L; Kolatsi-Joannou, Maria; Collinson, Sophie; Denny, Thomas; Knight, David; Mironov, Aleksandr; Starborg, Toby; Korstanje, Ron; Humphries, Martin J; Long, David A; Lennon, Rachel

    2015-12-01

    Glomerular disease often features altered histologic patterns of extracellular matrix (ECM). Despite this, the potential complexities of the glomerular ECM in both health and disease are poorly understood. To explore whether genetic background and sex determine glomerular ECM composition, we investigated two mouse strains, FVB and B6, using RNA microarrays of isolated glomeruli combined with proteomic glomerular ECM analyses. These studies, undertaken in healthy young adult animals, revealed unique strain- and sex-dependent glomerular ECM signatures, which correlated with variations in levels of albuminuria and known predisposition to progressive nephropathy. Among the variation, we observed changes in netrin 4, fibroblast growth factor 2, tenascin C, collagen 1, meprin 1-α, and meprin 1-β. Differences in protein abundance were validated by quantitative immunohistochemistry and Western blot analysis, and the collective differences were not explained by mutations in known ECM or glomerular disease genes. Within the distinct signatures, we discovered a core set of structural ECM proteins that form multiple protein-protein interactions and are conserved from mouse to man. Furthermore, we found striking ultrastructural changes in glomerular basement membranes in FVB mice. Pathway analysis of merged transcriptomic and proteomic datasets identified potential ECM regulatory pathways involving inhibition of matrix metalloproteases, liver X receptor/retinoid X receptor, nuclear factor erythroid 2-related factor 2, notch, and cyclin-dependent kinase 5. These pathways may therefore alter ECM and confer susceptibility to disease. PMID:25896609

  8. Extracellular Wound Matrices:A Novel Regenerative Tissue Matrix (RTM) Technology for Connective Tissue Reconstruction.

    PubMed

    Harper, John R; McQuillan, David J

    2007-06-01

    The restoration of structure, function, and physiology to damaged or missing tissue through the use of a regenerative tissue matrix (RTM) leads to regenerative healing rather than reparative scarring. While many processes exist to transform biologic materials into an extracellular matrix (ECM), only those that maintain the required structural and biochemical properties necessary to capture the intrinsic regenerative abilities of the body are suitable to produce an RTM. Histological examination using differential staining with hematoxylin and eosin stain or Verhoeff von Geisen stain of human biopsies of RTM obtained from 2 different abdominal surgery patients taken at 8- and 12 months were consistent with RTM remodeling into fascia-like tissue. A synopsis of recent studies on the use of the RTM GraftJacket® (Wright Medical Technologies, Memphis, Tenn) in successful closure of diabetic foot wounds is presented. Collectively, these reports indicate that LifeCell produced ECMs exemplified by GraftJacket exhibit the required clinical outcomes associated with an RTM. PMID:26110325

  9. The endogenous fluorescence of fibroblast in collagen gels as indicator of stiffness of the extracellular matrix

    NASA Astrophysics Data System (ADS)

    Padilla-Martinez, J. P.; Ortega-Martinez, A.; Franco, W.

    2016-03-01

    The stiffness or rigidity of the extracellular matrix (ECM) regulates cell response. Established mechanical tests to measure stiffness, such as indentation and tensile tests, are invasive and destructive to the sample. Endogenous or native molecules to cells and ECM components, like tryptophan and cross-links of collagen, display fluorescence upon irradiation with ultraviolet light. Most likely, the concentration of these endogenous fluorophores changes as the stiffness of the ECM changes. In this work we investigate the endogenous fluorescence of collagen gels containing fibroblasts as a non-invasive non-destructive method to measure stiffness of the ECM. Human fibroblast cells were cultured in three-dimensional gels of type I collagen (50,000 cells/ml). This construct is a simple model of tissue contraction. During contraction, changes in the excitation-emission matrix (a fluorescence map in the 240-520/290-530 nm range) of constructs were measured with a spectrofluoremeter, and changes in stiffness were measured with a standard indentation test over 16 days. Results show that a progressive increase in fluorescence of the 290/340 nm excitation-emission pair correlates with a progressive increase in stiffness (r=0.9, α=0.5). The fluorescence of this excitation-emission pair is ascribed to tryptophan and variations in the fluorescence of this pair correlate with cellular proliferation. In this tissue model, the endogenous functional fluorescence of proliferating fibroblast cells is a biomechanical marker of stiffness of the ECM.

  10. A Protein Involved in the Assembly of an Extracellular Calcium Storage Matrix*

    PubMed Central

    Glazer, Lilah; Shechter, Assaf; Tom, Moshe; Yudkovski, Yana; Weil, Simy; Aflalo, Eliahu David; Pamuru, Ramachandra Reddy; Khalaila, Isam; Bentov, Shmuel; Berman, Amir; Sagi, Amir

    2010-01-01

    Gastroliths, the calcium storage organs of crustaceans, consist of chitin-protein-mineral complexes in which the mineral component is stabilized amorphous calcium carbonate. To date, only three proteins, GAP 65, gastrolith matrix protein (GAMP), and orchestin, have been identified in gastroliths. Here, we report a novel protein, GAP 10, isolated from the gastrolith of the crayfish Cherax quadricarinatus and specifically expressed in its gastrolith disc. The encoding gene was cloned by partial sequencing of the protein extracted from the gastrolith matrix. Based on an assembled microarray cDNA chip, GAP 10 transcripts were found to be highly (12-fold) up-regulated in premolt gastrolith disc and significantly down-regulated in the hypodermis at the same molt stage. The deduced protein sequence of GAP 10 lacks chitin-binding domains and does not show homology to known proteins in the GenBankTM data base. It does, however, have an amino acid composition that has similarity to proteins extracted from invertebrate and ascidian-calcified extracellular matrices. The GAP 10 sequence contains a predicted signal peptide and predicted phosphorylation sites. In addition, the protein is phosphorylated and exhibits calcium-binding ability. Repeated daily injections of GAP 10 double strand RNA to premolt C. quadricarinatus resulted in a prolonged premolt stage and in the development of gastroliths with irregularly rough surfaces. These findings suggest that GAP 10 may be involved in the assembly of the gastrolith chitin-protein-mineral complex, particularly in the deposition of amorphous calcium carbonate. PMID:20150428

  11. A protein involved in the assembly of an extracellular calcium storage matrix.

    PubMed

    Glazer, Lilah; Shechter, Assaf; Tom, Moshe; Yudkovski, Yana; Weil, Simy; Aflalo, Eliahu David; Pamuru, Ramachandra Reddy; Khalaila, Isam; Bentov, Shmuel; Berman, Amir; Sagi, Amir

    2010-04-23

    Gastroliths, the calcium storage organs of crustaceans, consist of chitin-protein-mineral complexes in which the mineral component is stabilized amorphous calcium carbonate. To date, only three proteins, GAP 65, gastrolith matrix protein (GAMP), and orchestin, have been identified in gastroliths. Here, we report a novel protein, GAP 10, isolated from the gastrolith of the crayfish Cherax quadricarinatus and specifically expressed in its gastrolith disc. The encoding gene was cloned by partial sequencing of the protein extracted from the gastrolith matrix. Based on an assembled microarray cDNA chip, GAP 10 transcripts were found to be highly (12-fold) up-regulated in premolt gastrolith disc and significantly down-regulated in the hypodermis at the same molt stage. The deduced protein sequence of GAP 10 lacks chitin-binding domains and does not show homology to known proteins in the GenBank data base. It does, however, have an amino acid composition that has similarity to proteins extracted from invertebrate and ascidian-calcified extracellular matrices. The GAP 10 sequence contains a predicted signal peptide and predicted phosphorylation sites. In addition, the protein is phosphorylated and exhibits calcium-binding ability. Repeated daily injections of GAP 10 double strand RNA to premolt C. quadricarinatus resulted in a prolonged premolt stage and in the development of gastroliths with irregularly rough surfaces. These findings suggest that GAP 10 may be involved in the assembly of the gastrolith chitin-protein-mineral complex, particularly in the deposition of amorphous calcium carbonate. PMID:20150428

  12. Chimeric Aptamer-Gelatin Hydrogels as an Extracellular Matrix Mimic for Loading Cells and Growth Factors

    PubMed Central

    Zhang, Xiaolong; Battig, Mark R.; Chen, Niancao; Gaddes, Erin R.; Duncan, Katelyn L.; Wang, Yong

    2016-01-01

    It is important to synthesize materials to recapitulate critical functions of biological systems for a variety of applications such as tissue engineering and regenerative medicine. The purpose of this study was to synthesize a chimeric hydrogel as a promising extracellular matrix (ECM) mimic using gelatin, a nucleic acid aptamer and polyethylene glycol (PEG). This hydrogel had a macroporous structure that was highly permeable for fast molecular transport. Despite its high permeability, it could strongly sequester and sustainably release growth factors with high bioactivity. Notably, growth factors retained in the hydrogel could maintain ~50% bioactivity during a 14-day release test. It also provided cells with effective binding sites, which led to high efficiency of cell loading into the macroporous hydrogel matrix. When cells and growth factors were co-loaded into the chimeric hydrogel, living cells could still be observed by day 14 in a static serum-reduced culture condition. Thus, this chimeric aptamer-gelatin hydrogel constitutes a promising biomolecular ECM mimic for loading cells and growth factors. PMID:26791559

  13. Development of a Biological Scaffold Engineered Using the Extracellular Matrix Secreted by Skeletal Muscle Cells

    PubMed Central

    Hurd, Shiloh; Bhati, Nadia; Walker, Addison; Kasukonis, Ben; Wolchok, Jeffrey C.

    2015-01-01

    The performance of implantable biomaterials derived from decellularized tissue, including encouraging results with skeletal muscle, suggests that the extracellular matrix (ECM) derived from native tissue has promising regenerative potential. Yet, the supply of biomaterials derived from donated tissue will always be limited, which is why the in-vitro fabrication of ECM biomaterials that mimic the properties of tissue is an attractive alternative. Towards this end, our group has utilized a novel method to collect the ECM that skeletal muscle myoblasts secrete and form it into implantable scaffolds. The cell derived ECM contained several matrix constituents, including collagen and fibronectin that were also identified within skeletal muscle samples. The ECM was organized into a porous network that could be formed with the elongated and aligned architecture observed within muscle samples. The ECM material supported the attachment and in-vitro proliferation of cells, suggesting effectiveness for cell transplantation, and was well tolerated by the host when examined in-vivo. The results suggest that the ECM collection approach can be used to produce biomaterials with compositions and structures that are similar to muscle samples, and while the physical properties may not yet match muscle values, the in-vitro and in-vivo results indicate it may be a suitable first generation alternative to tissue derived biomaterials. PMID:25725550

  14. Study of extracellular matrix in vocal fold biomechanics using a two-phase model

    PubMed Central

    Li, Nicole Y. K.; Avazmohammadi, Reza; Thibeault, Susan L.; Mongrain, Rosaire; Mongeau, Luc

    2014-01-01

    The extracellular matrix (ECM) of the vocal fold tissue consists primarily of fibrous and interstitial proteins. The purpose of this study was to investigate the effects of selective enzymatic digestion of two ECM proteins, namely elastin and versican, on the elasticity of rabbit vocal fold tissue. Quasi-static, sinusoidal, uniaxial tensile tests were performed. The data were analyzed within the framework of a model of the ECM as a two-phase composite material consisting of collagen fibrils as the reinforcing fibers and noncollagenous ECM proteins as the matrix. To validate the two-phase model, the regression parameters for the fibers’ volume fraction and shear modulus in a different animal model were compared with corresponding published data. The proposed model was then used to analyze rabbit vocal fold tissues. The mean value and the standard deviation of the fiber volume fraction were found to be 8.49 ±3.75% for the control samples (n =4), 0.59 ±1.13 % after elastin removal (n =4), and 8.22 ±1.06% after versican removal (n =4). The results suggest that elastin removal may lead to a reduction in tissue stiffness, through counteracting the reinforcement of collagen fibrils. PMID:24792897

  15. Integrin-extracellular matrix interactions in connective tissue remodeling and osteoblast differentiation

    NASA Technical Reports Server (NTRS)

    Globus, R. K.; Moursi, A.; Zimmerman, D.; Lull, J.; Damsky, C.

    1995-01-01

    The differentiaton of bone cells is a complex multistep process. Bone is somewhat unusual in that it is very actively and continually remodeled in the adult and that maintenance of its mass in the mature organism is exquisitely sensitive to mechanical as well as chemical signals. Bone is also unique because it consists of a very large amount of extracellular matrix (ECM) that is mineralized. The integrin family of ECM receptors has been shown to play an important role in tissue morphogenesis in several systems. Our studies on the regulation of matrix remodeling enzymes by integrins in rabbit synovial fibroblasts show that two b1 integrin fibronectin (FN) receptor complexes (alpha 5 beta 1 and alpha 4 beta 1) cooperate in detecting subtle changes in the composition of the ECM. As a result of signal transduction by these integrins, the levels of mRNA and protein for several members of the metalloproteinase family are regulated in these cells. We have also used antibody and RGD peptide perturbation studies to determine the significance of cell/ECM interactions to normal osteogenesis. We found that interactions between the cell binding domain of FN and integrins are required for both normal morphogenesis and gene expression in cultured osteoblasts that differentiate to form bone-like tissue in culture. These data lead us to propose that beta 1 integrins play an important role in osteoblast differentiation as well as in bone remodeling.

  16. Role of the extracellular matrix protein thrombospondin in the early development of the mouse embryo

    PubMed Central

    1990-01-01

    The distribution of the extracellular matrix protein thrombospondin (TSP) in cleavage to egg cylinder staged mouse embryos and its role in trophoblast outgrowth from cultured blastocysts were examined. TSP was present within the cytoplasm of unfertilized eggs; in fertilized one- to four-cell embryos; by the eight-cell stage, TSP was also densely deposited at cell-cell borders. In the blastocyst, although TSP was present in all three cell types; trophectoderm, endoderm, and inner cell mass (ICM), it was enriched in the ICM and at the surface of trophectoderm cells. Hatched blastocysts grown on matrix-coated coverslips formed extensive trophoblast outgrowths on TSP, grew slightly less avidly on laminin, or on a 140-kD fragment of TSP containing its COOH terminus and putative cell binding domains. There was little outgrowth on the NH2 terminus heparin-binding domain. Addition of anti-TSP antibodies (but not GRGDS) to blastocysts growing on TSP strikingly inhibited outgrowth. Consistent with its early appearance and presence in trophoblast cells during implantation, TSP may play an important role in the early events involved in mammalian embryogenesis. PMID:2277082

  17. LKB1 loss in melanoma disrupts directional migration toward extracellular matrix cues

    PubMed Central

    Chan, Keefe T.; Asokan, Sreeja B.; King, Samantha J.; Bo, Tao; Dubose, Evan S.; Liu, Wenjin; Berginski, Matthew E.; Simon, Jeremy M.; Davis, Ian J.; Gomez, Shawn M.; Sharpless, Norman E.

    2014-01-01

    Somatic inactivation of the serine/threonine kinase gene STK11/LKB1/PAR-4 occurs in a variety of cancers, including ∼10% of melanoma. However, how the loss of LKB1 activity facilitates melanoma invasion and metastasis remains poorly understood. In LKB1-null cells derived from an autochthonous murine model of melanoma with activated Kras and Lkb1 loss and matched reconstituted controls, we have investigated the mechanism by which LKB1 loss increases melanoma invasive motility. Using a microfluidic gradient chamber system and time-lapse microscopy, in this paper, we uncover a new function for LKB1 as a directional migration sensor of gradients of extracellular matrix (haptotaxis) but not soluble growth factor cues (chemotaxis). Systematic perturbation of known LKB1 effectors demonstrated that this response does not require canonical adenosine monophosphate–activated protein kinase (AMPK) activity but instead requires the activity of the AMPK-related microtubule affinity-regulating kinase (MARK)/PAR-1 family kinases. Inhibition of the LKB1–MARK pathway facilitated invasive motility, suggesting that loss of the ability to sense inhibitory matrix cues may promote melanoma invasion. PMID:25349262

  18. Cellular and nerve regeneration within a biosynthetic extracellular matrix for corneal transplantation

    NASA Astrophysics Data System (ADS)

    Li, Fengfu; Carlsson, David; Lohmann, Chris; Suuronen, Erik; Vascotto, Sandy; Kobuch, Karin; Sheardown, Heather; Munger, Rejean; Nakamura, Masatsugu; Griffith, May

    2003-12-01

    Our objective was to determine whether key properties of extracellular matrix (ECM) macromolecules can be replicated within tissue-engineered biosynthetic matrices to influence cellular properties and behavior. To achieve this, hydrated collagen and N-isopropylacrylamide copolymer-based ECMs were fabricated and tested on a corneal model. The structural and immunological simplicity of the cornea and importance of its extensive innervation for optimal functioning makes it an ideal test model. In addition, corneal failure is a clinically significant problem. Matrices were therefore designed to have the optical clarity and the proper dimensions, curvature, and biomechanical properties for use as corneal tissue replacements in transplantation. In vitro studies demonstrated that grafting of the laminin adhesion pentapeptide motif, YIGSR, to the hydrogels promoted epithelial stratification and neurite in-growth. Implants into pigs' corneas demonstrated successful in vivo regeneration of host corneal epithelium, stroma, and nerves. In particular, functional nerves were observed to rapidly regenerate in implants. By comparison, nerve regeneration in allograft controls was too slow to be observed during the experimental period, consistent with the behavior of human cornea transplants. Other corneal substitutes have been produced and tested, but here we report an implantable matrix that performs as a physiologically functional tissue substitute and not simply as a prosthetic device. These biosynthetic ECM replacements should have applicability to many areas of tissue engineering and regenerative medicine, especially where nerve function is required. regenerative medicine | tissue engineering | cornea | implantation | innervation

  19. Extracellular matrix and growth factor engineering for controlled angiogenesis in regenerative medicine.

    SciTech Connect

    Martino, Mikael M.; Brkic, Sime; Bovo, Emmanuela; Burger, Maximilian; Schaefer, Dirk J.; Wolff, Thomas; Gurke, Lorenz; Briquez, Priscilla S.; Larsson, Hans M.; Gianni-Barrera, Roberto; Hubbell, Jeffrey A.; Banfi, Andrea

    2015-04-01

    Blood vessel growth plays a key role in regenerative medicine, both to restore blood supply to ischemic tissues and to ensure rapid vascularization of clinical-size tissue-engineered grafts. For example, vascular endothelial growth factor (VEGF) is the master regulator of physiological blood vessel growth and is one of the main molecular targets of therapeutic angiogenesis approaches. However, angiogenesis is a complex process and there is a need to develop rational therapeutic strategies based on a firm understanding of basic vascular biology principles, as evidenced by the disappointing results of initial clinical trials of angiogenic factor delivery. In particular, the spatial localization of angiogenic signals in the extracellular matrix (ECM) is crucial to ensure the proper assembly and maturation of new vascular structures. Here, we discuss the therapeutic implications of matrix interactions of angiogenic factors, with a special emphasis on VEGF, as well as provide an overview of current approaches, based on protein and biomaterial engineering that mimic the regulatory functions of ECM to optimize the signaling microenvironment of vascular growth factors.

  20. Extracellular matrix and growth factor engineering for controlled angiogenesis in regenerative medicine

    DOE PAGESBeta

    Martino, Mikael M.; Brkic, Sime; Bovo, Emmanuela; Burger, Maximilian; Schaefer, Dirk J.; Wolff, Thomas; Gurke, Lorenz; Briquez, Priscilla S.; Larsson, Hans M.; Gianni-Barrera, Roberto; et al

    2015-04-01

    In this study, blood vessel growth plays a key role in regenerative medicine, both to restore blood supply to ischemic tissues and to ensure rapid vascularization of clinical-size tissue-engineered grafts. For example, vascular endothelial growth factor (VEGF) is the master regulator of physiological blood vessel growth and is one of the main molecular targets of therapeutic angiogenesis approaches. However, angiogenesis is a complex process and there is a need to develop rational therapeutic strategies based on a firm understanding of basic vascular biology principles, as evidenced by the disappointing results of initial clinical trials of angiogenic factor delivery. In particular,more » the spatial localization of angiogenic signals in the extracellular matrix (ECM) is crucial to ensure the proper assembly and maturation of new vascular structures. Here, we discuss the therapeutic implications of matrix interactions of angiogenic factors, with a special emphasis on VEGF, as well as provide an overview of current approaches, based on protein and biomaterial engineering that mimic the regulatory functions of ECM to optimize the signaling microenvironment of vascular growth factors.« less

  1. Buckling Reduces eNOS Production and Stimulates Extracellular Matrix Remodeling in Arteries in Organ Culture.

    PubMed

    Xiao, Yangming; Liu, Qin; Han, Hai-Chao

    2016-09-01

    Artery buckling alters the fluid shear stress and wall stress in the artery but its temporal effect on vascular wall remodeling is poorly understood. The purpose of this study was to investigate the early effect of artery buckling on endothelial nitric oxide synthase (eNOS) expression and extracellular matrix remodeling. Bilateral porcine carotid arteries were maintained in an ex vivo organ culture system with and without buckling while under the same physiological pressure and flow rate for 3-7 days. Matrix metalloproteinase-2 (MMP-2), MMP-9, fibronectin, elastin, collagen I, III and IV, tissue inhibitor of metalloproteinase-2 (TIMP-2), and eNOS were determined using Western blotting and immunohistochemistry. Our results showed that MMP-2 expression level was significantly higher in buckled arteries than in the controls and higher at the inner curve than at the outer curve of buckled arteries, while collagen IV content showed an opposite trend, suggesting that artery buckling increased MMP-2 expression and collagen IV degradation in a site-specific fashion. However, no differences for MMP-9, fibronectin, elastin, collagen I, III, and TIMP-2 were observed among the outer and inner curve sides of buckled arteries and straight controls. Additionally, eNOS expression was significantly decreased in buckled arteries. These results suggest that artery buckling triggers uneven wall remodeling that could lead to development of tortuous arteries. PMID:26913855

  2. Periostin is an extracellular matrix protein required for eruption of incisors in mice

    SciTech Connect

    Kii, Isao; Amizuka, Norio; Minqi, Li; Kitajima, Satoshi; Saga, Yumiko; Kudo, Akira . E-mail: akudo@bio.titech.ac.jp

    2006-04-14

    A characteristic tooth of rodents, the incisor continuously grows throughout life by the constant formation of dentin and enamel. Continuous eruption of the incisor is accompanied with formation of shear zone, in which the periodontal ligament is remodeled. Although the shear zone plays a role in the remodeling, its molecular biological aspect is barely understood. Here, we show that periostin is essential for formation of the shear zone. Periostin {sup -/-} mice showed an eruption disturbance of incisors. Histological observation revealed that deletion of periostin led to disappearance of the shear zone. Electron microscopy revealed that the disappearance of the shear zone resulted from a failure in digestion of collagen fibers in the periostin {sup -/-} mice. Furthermore, immunohistochemical analysis using anti-periostin antibodies demonstrated the restricted localization of periostin protein in the shear zone. Periostin is an extracellular matrix protein, and immunoelectron microscopy showed a close association of periostin with collagen fibrils in vivo. These results suggest that periostin functions in the remodeling of collagen matrix in the shear zone.

  3. Extracellular matrix and growth factor engineering for controlled angiogenesis in regenerative medicine

    SciTech Connect

    Martino, Mikael M.; Brkic, Sime; Bovo, Emmanuela; Burger, Maximilian; Schaefer, Dirk J.; Wolff, Thomas; Gurke, Lorenz; Briquez, Priscilla S.; Larsson, Hans M.; Gianni-Barrera, Roberto; Hubbell, Jeffrey A.; Banfi, Andrea

    2015-04-01

    In this study, blood vessel growth plays a key role in regenerative medicine, both to restore blood supply to ischemic tissues and to ensure rapid vascularization of clinical-size tissue-engineered grafts. For example, vascular endothelial growth factor (VEGF) is the master regulator of physiological blood vessel growth and is one of the main molecular targets of therapeutic angiogenesis approaches. However, angiogenesis is a complex process and there is a need to develop rational therapeutic strategies based on a firm understanding of basic vascular biology principles, as evidenced by the disappointing results of initial clinical trials of angiogenic factor delivery. In particular, the spatial localization of angiogenic signals in the extracellular matrix (ECM) is crucial to ensure the proper assembly and maturation of new vascular structures. Here, we discuss the therapeutic implications of matrix interactions of angiogenic factors, with a special emphasis on VEGF, as well as provide an overview of current approaches, based on protein and biomaterial engineering that mimic the regulatory functions of ECM to optimize the signaling microenvironment of vascular growth factors.

  4. Cartilaginous extracellular matrix-modified chitosan hydrogels for cartilage tissue engineering.

    PubMed

    Choi, Bogyu; Kim, Soyon; Lin, Brian; Wu, Benjamin M; Lee, Min

    2014-11-26

    Cartilaginous extracellular matrix (ECM) components such as type-II collagen (Col II) and chondroitin sulfate (CS) play a crucial role in chondrogenesis. However, direct clinical use of natural Col II or CS as scaffolds for cartilage tissue engineering is limited by their instability and rapid enzymatic degradation. Here, we investigate the incorporation of Col II and CS into injectable chitosan hydrogels designed to gel upon initiation by exposure to visible blue light (VBL) in the presence of riboflavin. Unmodified chitosan hydrogel supported proliferation and deposition of cartilaginous ECM by encapsulated chondrocytes and mesenchymal stem cells. The incorporation of native Col II or CS into chitosan hydrogels further increased chondrogenesis. The incorporation of Col II, in particular, was found to be responsible for the enhanced cellular condensation and chondrogenesis observed in modified hydrogels. This was mediated by integrin α10 binding to Col II, increasing cell-matrix adhesion. These findings demonstrate the potential of cartilage ECM-modified chitosan hydrogels as biomaterials to promote cartilage regeneration. PMID:25361212

  5. Nrf2 null enhances UVB-induced skin inflammation and extracellular matrix damages.

    PubMed

    Saw, Constance Lay Lay; Yang, Anne Yuqing; Huang, Mou-Tuan; Liu, Yue; Lee, Jong Hun; Khor, Tin Oo; Su, Zheng-Yuan; Shu, Limin; Lu, Yaoping; Conney, Allan H; Kong, Ah-Ng Tony

    2014-01-01

    Nrf2 plays a critical role in defending against oxidative stress and inflammation. We previously reported that Nrf2 confers protection against ultraviolet-B (UVB)-induced inflammation, sunburn reaction, and is involved in sulforaphane-mediated photo-protective effects in the skin. In this study, we aimed to demonstrate the protective role of Nrf2 against inflammation-mediated extracellular matrix (ECM) damage induced by UVB irradiation. Ear biopsy weights were significantly increased in both Nrf2 wild-type (Nrf2 WT) and knockout (Nrf2 KO) mice one week after UVB irradiation. However, these weights increased more significantly in KO mice compared to WT mice, suggesting a greater inflammatory response in KO mice. In addition, we analyzed the protein expression of numerous markers, including macrophage inflammatory protein-2 (MIP-2), pro-matrix metalloproteinase-9 (MMP-9), and p53. p53, a regulator of DNA repair, was overexpressed in Nrf2 KO mice, indicating that the absence of Nrf2 led to more sustained DNA damage. There was also more substantial ECM degradation and increased inflammation in UVB-irradiated Nrf2 KO mice compared to UVB-irradiated WT mice. Furthermore, the protective effects of Nrf2 in response to UVB irradiation were mediated by increased HO-1 protein expression. Collectively, our results show that Nrf2 plays a key role in protecting against UVB irradiation and that the photo-protective effect of Nrf2 is closely related to the inhibition of ECM degradation and inflammation. PMID:25228981

  6. Fibers in the extracellular matrix enable long-range stress transmission between cells.

    PubMed

    Ma, Xiaoyue; Schickel, Maureen E; Stevenson, Mark D; Sarang-Sieminski, Alisha L; Gooch, Keith J; Ghadiali, Samir N; Hart, Richard T

    2013-04-01

    Cells can sense, signal, and organize via mechanical forces. The ability of cells to mechanically sense and respond to the presence of other cells over relatively long distances (e.g., ∼100 μm, or ∼10 cell-diameters) across extracellular matrix (ECM) has been attributed to the strain-hardening behavior of the ECM. In this study, we explore an alternative hypothesis: the fibrous nature of the ECM makes long-range stress transmission possible and provides an important mechanism for long-range cell-cell mechanical signaling. To test this hypothesis, confocal reflectance microscopy was used to develop image-based finite-element models of stress transmission within fibroblast-seeded collagen gels. Models that account for the gel's fibrous nature were compared with homogenous linear-elastic and strain-hardening models to investigate the mechanisms of stress propagation. Experimentally, cells were observed to compact the collagen gel and align collagen fibers between neighboring cells within 24 h. Finite-element analysis revealed that stresses generated by a centripetally contracting cell boundary are concentrated in the relatively stiff ECM fibers and are propagated farther in a fibrous matrix as compared to homogeneous linear elastic or strain-hardening materials. These results support the hypothesis that ECM fibers, especially aligned ones, play an important role in long-range stress transmission. PMID:23561517

  7. Study of extracellular matrix in vocal fold biomechanics using a two-phase model.

    PubMed

    Miri, Amir K; Li, Nicole Y K; Avazmohammadi, Reza; Thibeault, Susan L; Mongrain, Rosaire; Mongeau, Luc

    2015-01-01

    The extracellular matrix (ECM) of the vocal fold tissue consists primarily of fibrous and interstitial proteins. The purpose of this study was to investigate the effects of selective enzymatic digestion of two ECM proteins, namely elastin and versican, on the elasticity of rabbit vocal fold tissue. Quasi-static, sinusoidal, uniaxial tensile tests were performed. The data were analyzed within the framework of a model of the ECM as a two-phase composite material consisting of collagen fibrils as the reinforcing fibers and noncollagenous ECM proteins as the matrix. To validate the two-phase model, the regression parameters for the fibers' volume fraction and shear modulus in a different animal model were compared with corresponding published data. The proposed model was then used to analyze rabbit vocal fold tissues. The mean value and the standard deviation of the fiber volume fraction were found to be 8.49 ± 3.75 % for the control samples (n = 4), 0.59 ± 1.13 % after elastin removal (n = 4), and 8.22 ± 1.06 % after versican removal (n = 4). The results suggest that elastin removal may lead to a reduction in tissue stiffness, through counteracting the reinforcement of collagen fibrils. PMID:24792897

  8. Biomimetic Porous PLGA Scaffolds Incorporating Decellularized Extracellular Matrix for Kidney Tissue Regeneration.

    PubMed

    Lih, Eugene; Park, Ki Wan; Chun, So Young; Kim, Hyuncheol; Kwon, Tae Gyun; Joung, Yoon Ki; Han, Dong Keun

    2016-08-24

    Chronic kidney disease is now recognized as a major health problem, but current therapies including dialysis and renal replacement have many limitations. Consequently, biodegradable scaffolds to help repairing injured tissue are emerging as a promising approach in the field of kidney tissue engineering. Poly(lactic-co-glycolic acid) (PLGA) is a useful biomedical material, but its insufficient biocompatibility caused a reduction in cell behavior and function. In this work, we developed the kidney-derived extracellular matrix (ECM) incorporated PLGA scaffolds as a cell supporting material for kidney tissue regeneration. Biomimetic PLGA scaffolds (PLGA/ECM) with different ECM concentrations were prepared by an ice particle leaching method, and their physicochemical and mechanical properties were characterized through various analyses. The proliferation of renal cortical epithelial cells on the PLGA/ECM scaffolds increased with an increase in ECM concentrations (0.2, 1, 5, and 10%) in scaffolds. The PLGA scaffold containing 10% of ECM has been shown to be an effective matrix for the repair and reconstitution of glomerulus and blood vessels in partially nephrectomized mice in vivo, compared with only PLGA control. These results suggest that not only can the tissue-engineering techniques be an effective alternative method for treatment of kidney diseases, but also the ECM incorporated PLGA scaffolds could be promising materials for biomedical applications including tissue engineered scaffolds and biodegradable implants. PMID:27456613

  9. Sodium hydrosulfide prevents myocardial dysfunction through modulation of extracellular matrix accumulation and vascular density.

    PubMed

    Pan, Li-Long; Wang, Xian-Li; Wang, Xi-Ling; Zhu, Yi-Zhun

    2014-01-01

    The aim was to examine the role of exogenous hydrogen sulfide (H2S) on cardiac remodeling in post-myocardial infarction (MI) rats. MI was induced in rats by ligation of coronary artery. After treatment with sodium hydrosulfide (NaHS, an exogenous H2S donor, 56 μM/kg·day) for 42 days, the effects of NaHS on left ventricular morphometric features, echocardiographic parameters, heme oxygenase-1 (HO-1), matrix metalloproteinases-9 (MMP-9), type I and type III collagen, vascular endothelial growth factor (VEGF), CD34, and α-smooth muscle actin (α-SMA) in the border zone of infarct area were analyzed to elucidate the protective mechanisms of exogenous H2S on cardiac function and fibrosis. Forty-two days post MI, NaHS-treatment resulted in a decrease in myocardial fibrotic area in association with decreased levels of type I, type III collagen and MMP-9 and improved cardiac function. Meanwhile, NaHS administration significantly increased cystathionine γ-lyase (CSE), HO-1, α-SMA, and VEGF expression. This effect was accompanied by an increase in vascular density in the border zone of infarcted myocardium. Our results provided the strong evidences that exogenous H2S prevented cardiac remodeling, at least in part, through inhibition of extracellular matrix accumulation and increase in vascular density. PMID:25514418

  10. Fibers in the Extracellular Matrix Enable Long-Range Stress Transmission between Cells

    PubMed Central

    Ma, Xiaoyue; Schickel, Maureen E.; Stevenson, Mark D.; Sarang-Sieminski, Alisha L.; Gooch, Keith J.; Ghadiali, Samir N.; Hart, Richard T.

    2013-01-01

    Cells can sense, signal, and organize via mechanical forces. The ability of cells to mechanically sense and respond to the presence of other cells over relatively long distances (e.g., ∼100 μm, or ∼10 cell-diameters) across extracellular matrix (ECM) has been attributed to the strain-hardening behavior of the ECM. In this study, we explore an alternative hypothesis: the fibrous nature of the ECM makes long-range stress transmission possible and provides an important mechanism for long-range cell-cell mechanical signaling. To test this hypothesis, confocal reflectance microscopy was used to develop image-based finite-element models of stress transmission within fibroblast-seeded collagen gels. Models that account for the gel’s fibrous nature were compared with homogenous linear-elastic and strain-hardening models to investigate the mechanisms of stress propagation. Experimentally, cells were observed to compact the collagen gel and align collagen fibers between neighboring cells within 24 h. Finite-element analysis revealed that stresses generated by a centripetally contracting cell boundary are concentrated in the relatively stiff ECM fibers and are propagated farther in a fibrous matrix as compared to homogeneous linear elastic or strain-hardening materials. These results support the hypothesis that ECM fibers, especially aligned ones, play an important role in long-range stress transmission. PMID:23561517

  11. Activation of AMPK Prevents Monocrotaline-Induced Extracellular Matrix Remodeling of Pulmonary Artery

    PubMed Central

    Li, Shaojun; Han, Dong; Zhang, Yonghong; Xie, Xinming; Ke, Rui; Zhu, Yanting; Liu, Lu; Song, Yang; Yang, Lan; Li, Manxiang

    2016-01-01

    Background The current study was performed to investigate the effect of adenosine monophosphate (AMP) – activated protein kinase (AMPK) activation on the extracellular matrix (ECM) remodeling of pulmonary arteries in pulmonary arterial hypertension (PAH) and to address its potential mechanisms. Material/Methods PAH was induced by a single intraperitoneal injection of monocrotaline (MCT) into Sprague-Dawley rats. Metformin (MET) was administered to activate AMPK. Immunoblotting was used to determine the phosphorylation and expression of AMPK and expression of tissue inhibitor of metalloproteinase-1 (TIMP-1). Gelatin zymography was performed to determine the activity of matrix metalloproteinase-2 (MMP-2) and MMP-9. Results Activation of AMPK by MET significantly reduced the right ventricle systolic pressure and the right ventricular hypertrophy in MCT-induced rat PAH model, and partially inhibited the ECM remodeling of pulmonary arteries. These effects were coupled with the decrease of MMP-2/9 activity and TIMP-1 expression. Conclusions This study suggests that activation of AMPK benefits PAH by inhibiting ECM remodeling of pulmonary arteries. Enhancing AMPK activity might have potential value in clinical treatment of PAH. PMID:26978596

  12. Abnormal osteopontin and matrix extracellular phosphoglycoprotein localization, and odontoblast differentiation, in X-linked hypophosphatemic teeth.

    PubMed

    Salmon, B; Bardet, C; Coyac, B R; Baroukh, B; Naji, J; Rowe, P S; Opsahl Vital, S; Linglart, A; Mckee, M D; Chaussain, C

    2014-08-01

    Mutations in phosphate-regulating gene (PHEX) lead to X-linked hypophosphatemic rickets (XLH), a genetic disease characterized by impaired mineralization in bones and teeth. In human XLH tooth dentin, calcospherites that would normally merge as part of the mineralization process are separated by unmineralized interglobular spaces where fragments of matrix proteins accumulate. Here, we immunolocalized osteopontin (OPN) in human XLH teeth, in a three-dimensional XLH human dental pulp stem cell-collagen scaffold culture model and in a rat tooth injury repair model treated with acidic serine- and aspartate-rich motif peptides (ASARM). In parallel, matrix extracellular phosphoglycoprotein (MEPE) immunolocalization and alkaline phosphatase (ALP) activity were assessed in XLH teeth. OPN was expressed by odontoblasts in the XLH models, and localized to the abnormal calcospherites of XLH tooth dentin. In addition, ALP activity and MEPE localization were abnormal in human XLH teeth, with MEPE showing an accumulation in the unmineralized interglobular spaces in dentin. Furthermore, XLH odontoblasts failed to form a well-polarized odontoblast layer. These data suggest that both MEPE and OPN are involved in impaired tooth mineralization associated with XLH, possibly through different effects on the mineralization process. PMID:25158186

  13. Extracellular matrix alterations in experimental Leishmania amazonensis infection in susceptible and resistant mice

    PubMed Central

    2012-01-01

    Leishmania is inoculated, by the bite of an infected sandfly, into the skin of the host, where the promastigotes are phagocyted by dermal macrophages. The dermal region comprises cells and abundant extracellular matrix. Studies show that matrix metalloproteinases play an important role in host defense responses against pathogens in mammals and that their activities lead to the production of antimicrobial peptides. The aim of this study is to evaluate the changes in the distribution of fibronectin and laminin as well as in the elastic system fibres during the course of infection caused by Leishmania amazonensis in mice with distinct genetic backgrounds of susceptibility to this parasite. The results showed that BALB/c presented an enhancement of fibronectin during the course of infection when compared to their control group while the infected or non-infected C3H.He showed a decrease of this protein at end of the experiment. Laminin, on the other hand, remained unaltered in both strains. Also in both BALB/c and C3H.He mice the elastic and elaunin fibres remained unchanged while the oxytalan fibres decreased along the experiment. Ninety days after the infection C3H.He mice had recovered their capacity to produce oxytalan fibres. PMID:22316002

  14. Revealing cytokine-induced changes in the extracellular matrix with secondary ion mass spectrometry

    PubMed Central

    Taylor, Adam J; Ratner, Buddy D; Buttery, Lee DK; Alexander, Morgan R

    2015-01-01

    Cell-secreted matrices (CSMs), where extracellular matrix (ECM) deposited by monolayer cell cultures are decellularized, have been increasingly used to produce surfaces that may be reseeded with cells. Such surfaces are useful to help us understand cell-ECM interactions in a microenvironment closer to the in vivo situation than synthetic substrates with adsorbed proteins. We describe the production of CSMs from mouse primary osteoblasts (mPObs) exposed to cytokine challenge during matrix secretion, mimicking in vivo inflammatory environments. Time-of-flight secondary ion mass spectrometry (ToF-SIMS) data revealed that CSMs with cytokine challenge at day 7 or day 12 of culture can be chemically distinguished from one another and from untreated CSM using multivariate analysis. Comparison of the differences with reference spectra from adsorbed protein mixtures points towards cytokine challenge resulting in a decrease in collagen content. This is supported by immunocytochemical and histological staining, demonstrating a 44% loss of collagen mass and a 32% loss in collagen I coverage. CSM surfaces demonstrate greater cell adhesion than adsorbed ECM proteins. When mPObs were reseeded onto cytokine-challenged CSMs they exhibited reduced adhesion and elongated morphology compared to untreated CSMs. Such changes may direct subsequent cell fate and function and provide insights into pathological responses at sites of inflammation. PMID:25523877

  15. Chitinases and Imaginal disc growth factors organize the extracellular matrix formation at barrier tissues in insects

    PubMed Central

    Pesch, Yanina-Yasmin; Riedel, Dietmar; Patil, Kapil R; Loch, Gerrit; Behr, Matthias

    2016-01-01

    The cuticle forms an apical extracellular-matrix (ECM) that covers exposed organs, such as epidermis, trachea and gut, for organizing morphogenesis and protection of insects. Recently, we reported that cuticle proteins and chitin are involved in ECM formation. However, molecular mechanisms that control assembly, maturation and replacement of the ECM and its components are not well known. Here we investigated the poorly described glyco-18-domain hydrolase family in Drosophila and identified the Chitinases (Chts) and imaginal-disc-growth-factors (Idgfs) that are essential for larval and adult molting. We demonstrate that Cht and idgf depletion results in deformed cuticles, larval and adult molting defects, and insufficient protection against wounding and bacterial infection, which altogether leads to early lethality. We show that Cht2/Cht5/Cht7/Cht9/Cht12 and idgf1/idgf3/idgf4/idgf5/idgf6 are needed for organizing proteins and chitin-matrix at the apical cell surface. Our data indicate that normal ECM formation requires Chts, which potentially hydrolyze chitin-polymers. We further suggest that the non-enzymatic idgfs act as structural proteins to maintain the ECM scaffold against chitinolytic degradation. Conservation of Chts and Idgfs proposes analogous roles in ECM dynamics across the insect taxa, indicating that Chts/Idgfs are new targets for species specific pest control. PMID:26838602

  16. Extracellular matrix signatures of human mammary carcinoma identify novel metastasis promoters

    PubMed Central

    Naba, Alexandra; Clauser, Karl R; Lamar, John M; Carr, Steven A; Hynes, Richard O

    2014-01-01

    The extracellular matrix (ECM) is a major component of tumors and a significant contributor to cancer progression. In this study, we use proteomics to investigate the ECM of human mammary carcinoma xenografts and show that primary tumors of differing metastatic potential differ in ECM composition. Both tumor cells and stromal cells contribute to the tumor matrix and tumors of differing metastatic ability differ in both tumor- and stroma-derived ECM components. We define ECM signatures of poorly and highly metastatic mammary carcinomas and these signatures reveal up-regulation of signaling pathways including TGFβ and VEGF. We further demonstrate that several proteins characteristic of highly metastatic tumors (LTBP3, SNED1, EGLN1, and S100A2) play causal roles in metastasis, albeit at different steps. Finally we show that high expression of LTBP3 and SNED1 correlates with poor outcome for ER−/PR−breast cancer patients. This study thus identifies novel biomarkers that may serve as prognostic and diagnostic tools. DOI: http://dx.doi.org/10.7554/eLife.01308.001 PMID:24618895

  17. Decellularization of intact tissue enables MALDI imaging mass spectrometry analysis of the extracellular matrix.

    PubMed

    Gessel, Megan; Spraggins, Jeffrey M; Voziyan, Paul; Hudson, Billy G; Caprioli, Richard M

    2015-11-01

    Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is a powerful molecular mapping technology that offers unbiased visualization of the spatial arrangement of biomolecules in tissue. Although there has been a significant increase in the number of applications employing this technology, the extracellular matrix (ECM) has received little attention, likely because ECM proteins are mostly large, insoluble and heavily cross-linked. We have developed a new sample preparation approach to enable MALDI IMS analysis of ECM proteins in tissue. Prior to freezing and sectioning, intact tissues are decellularized by incubation in sodium dodecyl sulfate. Decellularization removes the highly abundant, soluble species that dominate a MALDI IMS spectrum while preserving the structural integrity of the ECM. In situ tryptic hydrolysis and imaging of tryptic peptides are then carried out to accommodate the large sizes of ECM proteins. This new approach allows the use of MALDI IMS for identification of spatially specific changes in ECM protein expression and modification in tissue. PMID:26505774

  18. [Theodor Huzella and the initiation of research on the interactions between cells and the extracellular matrix].

    PubMed

    Robert, Ladislas; Labat-Robert, Jacqueline; Michel Robert, Alexandre

    2012-01-01

    Interactions between cells and the surrounding "biomatrix", mediated by receptors as integrins or the elastin receptor is the most important topic in up to date research on connective tissues. Looking for the origin of this concept, one finds the pioneering work of Theodor Huzella, professor of histology-embryology at the Medical University of Budapest during pre-world war II decades. Using time-laps micro-cinematography in reflected light, he visualized the important role of connective tissue fibers, prepared in his laboratory, for the oriented migration of normal and malignant cells. His theoretical explanations, attributing an "active" role to the elasticity of the argyrophilic fibrous network in the coordination of cell societies, can now be reinterpreted in the light of recent work on the mechanotransduction of "messages" from the extracellular matrix to the cell inside. We propose a succinct review of Huzella's work and theories reinterpreted in the light of up-to-date knowledge on cell-matrix interactions. PMID:22748046

  19. Proliferation of progeria cells is enhanced by lamina-associated polypeptide 2α (LAP2α) through expression of extracellular matrix proteins

    PubMed Central

    Vidak, Sandra; Kubben, Nard; Dechat, Thomas; Foisner, Roland

    2015-01-01

    Lamina-associated polypeptide 2α (LAP2α) localizes throughout the nucleoplasm and interacts with the fraction of lamins A/C that is not associated with the peripheral nuclear lamina. The LAP2α–lamin A/C complex negatively affects cell proliferation. Lamins A/C are encoded by LMNA, a single heterozygous mutation of which causes Hutchinson-Gilford progeria syndrome (HGPS). This mutation generates the lamin A variant progerin, which we show here leads to loss of LAP2α and nucleoplasmic lamins A/C, impaired proliferation, and down-regulation of extracellular matrix components. Surprisingly, contrary to wild-type cells, ectopic expression of LAP2α in cells expressing progerin restores proliferation and extracellular matrix expression but not the levels of nucleoplasmic lamins A/C. We conclude that, in addition to its cell cycle-inhibiting function with lamins A/C, LAP2α can also regulate extracellular matrix components independently of lamins A/C, which may help explain the proliferation-promoting function of LAP2α in cells expressing progerin. PMID:26443848

  20. Proliferation of progeria cells is enhanced by lamina-associated polypeptide 2α (LAP2α) through expression of extracellular matrix proteins.

    PubMed

    Vidak, Sandra; Kubben, Nard; Dechat, Thomas; Foisner, Roland

    2015-10-01

    Lamina-associated polypeptide 2α (LAP2α) localizes throughout the nucleoplasm and interacts with the fraction of lamins A/C that is not associated with the peripheral nuclear lamina. The LAP2α-lamin A/C complex negatively affects cell proliferation. Lamins A/C are encoded by LMNA, a single heterozygous mutation of which causes Hutchinson-Gilford progeria syndrome (HGPS). This mutation generates the lamin A variant progerin, which we show here leads to loss of LAP2α and nucleoplasmic lamins A/C, impaired proliferation, and down-regulation of extracellular matrix components. Surprisingly, contrary to wild-type cells, ectopic expression of LAP2α in cells expressing progerin restores proliferation and extracellular matrix expression but not the levels of nucleoplasmic lamins A/C. We conclude that, in addition to its cell cycle-inhibiting function with lamins A/C, LAP2α can also regulate extracellular matrix components independently of lamins A/C, which may help explain the proliferation-promoting function of LAP2α in cells expressing progerin. PMID:26443848

  1. Constructing Human Skin Equivalents on Porcine Acellular Peritoneum Extracellular Matrix for In Vitro Irritation Testing.

    PubMed

    Tsai, Pei-Chin; Zhang, Zheng; Florek, Charles; Michniak-Kohn, Bozena B

    2016-01-01

    The irritancy of topical products has to be investigated to ensure the safety and compliance. Although several reconstructed human epidermal models have been adopted by the Organization for Economic Cooperation and Development (OECD) to replace in vivo animal irritation testing, these models are based on a single cell type and lack dermal components, which may be insufficient to reflect all of the components of irritation. In our study, we investigated the use of acellular porcine peritoneum extracellular matrix as a substrate to construct full-thickness human skin equivalents (HSEs) for use as irritation screening tool. The acellular peritoneum matrix (APM) exhibited excellent skin cell attachment (>80%) and proliferation for human dermal fibroblasts (HDF) and immortalized human keratinocytes (HaCaT). APM-HSEs based on coculture of HDF and HaCaT were prepared. Increased HDF seeding density up to 5 × 10(4)/cm(2) resulted in APM-HSEs with a thicker and more organized epidermis. The epidermis of APM-HSEs expressed keratin 15, a keratinocyte proliferation marker, and involucrin, a differentiation marker, respectively. To assess the use of APM-HSEs for irritation testing, six proficiency chemicals, including three nonirritants (phosphate-buffered saline, polyethylene glycol 400, and isopropanol) and three irritants (1-bromohexane, heptanol, and sodium dodecyl sulfate) were applied. The APM-HSEs were able to discriminate nonirritants from irritants based on the viability. Levels of cytokines (interleukin [IL]-1α, IL-1ra, IL-6, IL-8, and granulocyte macrophage colony-stimulating factor [GM-CSF]) in these treatment groups further assisted the irritancy ranking. In conclusion, we have developed partially differentiated full-thickness APM-HSEs based on acellular porcine peritoneum matrix, and these APM-HSEs demonstrated utility as an in vitro irritation screening tool. PMID:26415037

  2. Longitudinal Measurement of Extracellular Matrix Rigidity in 3D Tumor Models Using Particle-tracking Microrheology

    PubMed Central

    El-Hamidi, Hamid; Celli, Jonathan P.

    2014-01-01

    The mechanical microenvironment has been shown to act as a crucial regulator of tumor growth behavior and signaling, which is itself remodeled and modified as part of a set of complex, two-way mechanosensitive interactions. While the development of biologically-relevant 3D tumor models have facilitated mechanistic studies on the impact of matrix rheology on tumor growth, the inverse problem of mapping changes in the mechanical environment induced by tumors remains challenging. Here, we describe the implementation of particle-tracking microrheology (PTM) in conjunction with 3D models of pancreatic cancer as part of a robust and viable approach for longitudinally monitoring physical changes in the tumor microenvironment, in situ. The methodology described here integrates a system of preparing in vitro 3D models embedded in a model extracellular matrix (ECM) scaffold of Type I collagen with fluorescently labeled probes uniformly distributed for position- and time-dependent microrheology measurements throughout the specimen. In vitro tumors are plated and probed in parallel conditions using multiwell imaging plates. Drawing on established methods, videos of tracer probe movements are transformed via the Generalized Stokes Einstein Relation (GSER) to report the complex frequency-dependent viscoelastic shear modulus, G*(ω). Because this approach is imaging-based, mechanical characterization is also mapped onto large transmitted-light spatial fields to simultaneously report qualitative changes in 3D tumor size and phenotype. Representative results showing contrasting mechanical response in sub-regions associated with localized invasion-induced matrix degradation as well as system calibration, validation data are presented. Undesirable outcomes from common experimental errors and troubleshooting of these issues are also presented. The 96-well 3D culture plating format implemented in this protocol is conducive to correlation of microrheology measurements with therapeutic

  3. Extracellular Matrix Peptides of Artemia Cyst Shell Participate in Protecting Encysted Embryos from Extreme Environments

    PubMed Central

    Dai, Li; Chen, Dian-Fu; Liu, Yu-Lei; Zhao, Yang; Yang, Fan; Yang, Jin-Shu; Yang, Wei-Jun

    2011-01-01

    Background Many species of the brine shrimp Artemia are found in various severe environments in many parts of the world where extreme salinity, high UV radiation levels, high pH, anoxia, large temperature fluctuations, and intermittent dry conditions are often recorded. To withstand adverse environments, Artemia undergoes an oviparous developmental pathway to release cysts whereas, under favorable conditions, swimming nauplius larvae are formed directly via an ovoviviparous pathway. In the former case these cysts have an extraordinary ability to keep the embryos protected from the harsh environment for long periods. This is achieved through the protection by a complex out-wrapping cyst shell. However, the formation and function of the cyst shell is complex; the details remain largely unclear. Principal Finding A shell gland-specific gene (SGEG2) was cloned and identified from a suppression subtractive hybridization library. Western blot analysis showed that SGEG2 presumably requires post-translational proteolysis in order to be processed into two mature peptides (SGEG2a and 2b). The three matrix peptides (SGEG1 reported previously, 2a, and 2b) were found to distribute throughout the cyst shell. The results of gene knockdown by RNAi and subsequent resistance to environmental stresses assays indicated that these matrix peptides are required for cyst shell formation and are involved in protecting the encysted embryos from environmental stress. Conclusions/Significance This study revealed that extracellular matrix peptides participate in protecting embryos from extreme salinity, UV radiation, large temperature fluctuations and dry environments, thereby facilitating their survival. The cyst shell provides an excellent opportunity to link the ecological setting of an organism to the underlying physiological and biochemical processes enabling its survival. The cyst shell material has also a high potential to become an excellent new biomaterial with a high number of

  4. Extracellular matrix remodeling of the testes through the male reproductive cycle in Teleostei fish.

    PubMed

    Santana, Julio Cesar de Oliveira; Quagio-Grassiotto, Irani

    2014-12-01

    During the fish reproductive cycle, testes undergo morphological changes related to germinal epithelium and remodeling of extracellular matrix components (ECM). ECM is degraded mainly by action of matrix metalloproteinases (MMPs). Due to the natural renewal of ECM in fish testes, we choose Pimelodus maculatus to study remodeling of ECM throughout reproductive cycle, using picrosirius (to identify type I, II, III collagen) and reticulin (type III collagen), and to immunolocalize MT1-MMP (membrane type 1-matrix metalloproteinase) and MMP-2 in testis cells. Testes were classified in four reproductive phases: regenerating, development, spawning capable and regressing. Picrosirius and reticulin demonstrated a differential distribution of total collagen fibers during the reproductive cycle. Immunohistochemistry showed MT1-MMP only in acidophilic granulocyte cells mainly inside blood vessels, in connective tissue of capsule close to the germinal compartment, and also infiltrated in interstitial connective tissue. MMP-2 was detected in fibroblast and endothelial cells of interstitial and capsule blood vessels, in epithelial cells of capsule, and in acidophilic granulocyte cells at same description for MT1-MMP. The fish testes ECM were remodeled throughout reproductive cycle in according to morphophysiological alterations. During reproductive season (spawning capable), the interstitium increased in total collagen fibers (type I, II, III). After spermiation period (regression and regenerating), the amount of collagen fibers decreased in response to action of MMPs on collagen degradation and other interstitial components (not assessed in this study). MMPs seem to be indispensable components for natural cyclic events of ECM remodeling of fish testes and for guarantee tissue homeostasis throughout reproductive cycle. PMID:25142725

  5. Identification of a fibronectin interaction site in the extracellular matrix protein ameloblastin.

    PubMed

    Beyeler, Michael; Schild, Christof; Lutz, Roman; Chiquet, Matthias; Trueb, Beat

    2010-04-15

    Mammalian teeth are composed of hydroxyapatite crystals that are embedded in a rich extracellular matrix. This matrix is produced by only two cell types, the mesenchymal odontoblasts and the ectodermal ameloblasts. Ameloblasts secrete the enamel proteins amelogenin, ameloblastin, enamelin and amelotin. Odontoblasts secrete collagen type I and several calcium-binding phosphoproteins including dentin sialophosphoprotein, dentin matrix protein, bone sialoprotein and osteopontin. The latter four proteins have recently been grouped in the family of the SIBLINGs (small integrin-binding ligand, N-linked glycoproteins) because they display similar gene structures and because they contain an RGD tripeptide sequence that binds to integrin receptors and thus mediates cell adhesion. We have prepared all the other tooth-specific proteins in recombinant form and examined whether they might also promote cell adhesion similar to the SIBLINGs. We found that only ameloblastin consistently mediated adhesion of osteoblastic and fibroblastic cells to plastic or titanium surfaces. The activity was dependent on the intact three-dimensional structure of ameloblastin and required de novo protein synthesis of the adhering cells. By deletion analysis and in vitro mutagenesis, the active site could be narrowed down to a sequence of 13 amino acid residues (VPIMDFADPQFPT) derived from exon 7 of the rat ameloblastin gene or exons 7-9 of the human gene. Kinetic studies and RNA interference experiments further demonstrated that this sequence does not directly bind to a cell surface receptor but that it interacts with cellular fibronectin, which in turn binds to integrin receptors. The identification of a fibronectin-binding domain in ameloblastin might permit interesting applications for dental implantology. Implants could be coated with peptides containing the active sequence, which in turn would recruit fibronectin from the patient's blood. The recruited fibronectin should then promote cell

  6. Cardiomyoblast (H9c2) Differentiation on Tunable Extracellular Matrix Microenvironment

    PubMed Central

    Suhaeri, Muhammad; Subbiah, Ramesh; Van, Se Young; Du, Ping; Kim, In Gul; Lee, Kangwon

    2015-01-01

    Extracellular matrices (ECM) obtained from in vitro-cultured cells have been given much attention, but its application in cardiac tissue engineering is still limited. This study investigates cardiomyogenic potential of fibroblast-derived matrix (FDM) as a novel ECM platform over gelatin or fibronectin, in generating cardiac cell lineages derived from H9c2 cardiomyoblasts. As characterized through SEM and AFM, FDM exhibits unique surface texture and biomechanical property. Immunofluorescence also found fibronectin, collagen, and laminin in the FDM. Cells on FDM showed a more circular shape and slightly less proliferation in a growth medium. After being cultured in a differentiation medium for 7 days, H9c2 cells on FDM differentiated into cardiomyocytes, as identified by stronger positive markers, such as α-actinin and cTnT, along with more elevated gene expression of Myl2 and Tnnt compared to the cells on gelatin and fibronectin. The gap junction protein connexin 43 was also significantly upregulated for the cells differentiated on FDM. A successive work enabled matrix stiffness tunable; FDM crosslinked by 2wt% genipin increased the stiffness up to 8.5 kPa, 100 times harder than that of natural FDM. The gene expression of integrin subunit α5 was significantly more upregulated on FDM than on crosslinked FDM (X-FDM), whereas no difference was observed for β1 expression. Interestingly, X-FDM showed a much greater effect on the cardiomyoblast differentiation into cardiomyocytes over natural one. This study strongly indicates that FDM can be a favorable ECM microenvironment for cardiomyogenesis of H9c2 and that tunable mechanical compliance induced by crosslinking further provides a valuable insight into the role of matrix stiffness on cardiomyogenesis. PMID:25836924

  7. Renal extracellular matrix accumulation in acute puromycin aminonucleoside nephrosis in rats.

    PubMed Central

    Jones, C. L.; Buch, S.; Post, M.; McCulloch, L.; Liu, E.; Eddy, A. A.

    1992-01-01

    Progressive renal fibrosis is considered to be the final common pathway leading to chronic renal insufficiency. In this study, the authors examined some of the cellular and molecular mechanisms regulating the renal accumulation of extracellular matrix (ECM) proteins using rats with puromycin amino-nucleoside (PAN) nephrosis as an acute model system. Puromycin aminonucleoside rats developed reversible nephrotic syndrome accompanied by an interstitial infiltrate of monocytes. The number of interstitial fibroblasts expressing ST4 antigen did not increase. During the first 4 days, steady-state mRNA levels for all genes examined remained at or below control levels. At 1 week, nephrotic syndrome and interstitial inflammation were established, and a period of renal cell proliferation occurred, identified by increased histone mRNA levels and localized by tritiated thymine autoradiography to tubular epithelial cells and occasional interstitial cells. Transforming growth factor-beta (TGF-beta) steady-state mRNA levels were increased eightfold, but returned to control levels by 3 weeks. At week 1, there was a 10- to 20-fold increase in kidney steady-state mRNA levels for genes encoding interstitial matrix proteins collagen I and fibronectin and basement membrane collagen IV. By in situ hybridization, alpha 1(I) procollagen mRNA was localized to interstitial cells. Immunofluorescence microscopy demonstrated focal accumulation of ECM proteins in the tubulointerstitial compartment at 2 and 3 weeks, but by 6 weeks, kidney immunohistology was normal again. Steady-state mRNA levels for the matrix degrading metalloproteinase stromelysin remained at control values, whereas the levels for interstitial collagenase were normal at week 1 and increased twofold to threefold at 2 and 3 weeks. Steady-state mRNA levels for the tissue inhibitor of metalloproteinases (TIMP) increased fivefold at 1 week and returned to baseline values over the next 2 weeks. The results of this study suggest that

  8. Modulation of canonical Wnt signaling by the extracellular matrix component biglycan

    PubMed Central

    Berendsen, Agnes D.; Fisher, Larry W.; Kilts, Tina M.; Owens, Rick T.; Robey, Pamela G.; Gutkind, J. Silvio; Young, Marian F.

    2011-01-01

    Although extracellular control of canonical Wnt signaling is crucial for tissue homeostasis, the role of the extracellular microenvironment in modulating this signaling pathway is largely unknown. In the present study, we show that a member of the small leucine-rich proteoglycan family, biglycan, enhances canonical Wnt signaling by mediating Wnt function via its core protein. Immunoprecipitation analysis revealed that biglycan interacts with both the canonical Wnt ligand Wnt3a and the Wnt coreceptor low-density lipoprotein receptor-related protein 6 (LRP6), possibly via the formation of a trimeric complex. Biglycan-deficient cells treated with exogenous Wnt3a had less Wnt3a retained in cell layers compared with WT cells. Furthermore, the Wnt-induced levels of LRP6 phosphorylation and expression of several Wnt target genes were blunted in biglycan-deficient cells. Both recombinant biglycan proteoglycan and biglycan core protein increased Wnt-induced β-catenin/T cell-specific factor–mediated transcriptional activity, and this activity was completely inhibited by Dickkopf 1. Interestingly, recombinant biglycan was able to rescue impaired Wnt signaling caused by a previously described missense mutation in the extracellular domain of human LRP6 (R611C). Furthermore, biglycan's modulation of canonical Wnt signaling affected the functional activities of osteoprogenitor cells, including the RUNX2-mediated transcriptional activity and calcium deposition. Use of a transplant system and a fracture healing model revealed that expression of Wnt-induced secreted protein 1 was decreased in bone formed by biglycan-deficient cells, further suggesting reduced Wnt signaling in vivo. We propose that biglycan may serve as a reservoir for Wnt in the pericellular space and modulate Wnt availability for activation of the canonical Wnt pathway. PMID:21969569

  9. Fibroblast-Derived Extracellular Matrix Induces Chondrogenic Differentiation in Human Adipose-Derived Mesenchymal Stromal/Stem Cells in Vitro

    PubMed Central

    Dzobo, Kevin; Turnley, Taegyn; Wishart, Andrew; Rowe, Arielle; Kallmeyer, Karlien; van Vollenstee, Fiona A.; Thomford, Nicholas E.; Dandara, Collet; Chopera, Denis; Pepper, Michael S.; Parker, M. Iqbal

    2016-01-01

    Mesenchymal stromal/stem cells (MSCs) represent an area being intensively researched for tissue engineering and regenerative medicine applications. MSCs may provide the opportunity to treat diseases and injuries that currently have limited therapeutic options, as well as enhance present strategies for tissue repair. The cellular environment has a significant role in cellular development and differentiation through cell–matrix interactions. The aim of this study was to investigate the behavior of adipose-derived MSCs (ad-MSCs) in the context of a cell-derived matrix so as to model the in vivo physiological microenvironment. The fibroblast-derived extracellular matrix (fd-ECM) did not affect ad-MSC morphology, but reduced ad-MSC proliferation. Ad-MSCs cultured on fd-ECM displayed decreased expression of integrins α2 and β1 and subsequently lost their multipotency over time, as shown by the decrease in CD44, Octamer-binding transcription factor 4 (OCT4), SOX2, and NANOG gene expression. The fd-ECM induced chondrogenic differentiation in ad-MSCs compared to control ad-MSCs. Loss of function studies, through the use of siRNA and a mutant Notch1 construct, revealed that ECM-mediated ad-MSCs chondrogenesis requires Notch1 and β-catenin signaling. The fd-ECM also showed anti-senescence effects on ad-MSCs. The fd-ECM is a promising approach for inducing chondrogenesis in ad-MSCs and chondrogenic differentiated ad-MSCs could be used in stem cell therapy procedures. PMID:27527147

  10. Fibroblast-Derived Extracellular Matrix Induces Chondrogenic Differentiation in Human Adipose-Derived Mesenchymal Stromal/Stem Cells in Vitro.

    PubMed

    Dzobo, Kevin; Turnley, Taegyn; Wishart, Andrew; Rowe, Arielle; Kallmeyer, Karlien; van Vollenstee, Fiona A; Thomford, Nicholas E; Dandara, Collet; Chopera, Denis; Pepper, Michael S; Parker, M Iqbal

    2016-01-01

    Mesenchymal stromal/stem cells (MSCs) represent an area being intensively researched for tissue engineering and regenerative medicine applications. MSCs may provide the opportunity to treat diseases and injuries that currently have limited therapeutic options, as well as enhance present strategies for tissue repair. The cellular environment has a significant role in cellular development and differentiation through cell-matrix interactions. The aim of this study was to investigate the behavior of adipose-derived MSCs (ad-MSCs) in the context of a cell-derived matrix so as to model the in vivo physiological microenvironment. The fibroblast-derived extracellular matrix (fd-ECM) did not affect ad-MSC morphology, but reduced ad-MSC proliferation. Ad-MSCs cultured on fd-ECM displayed decreased expression of integrins α2 and β1 and subsequently lost their multipotency over time, as shown by the decrease in CD44, Octamer-binding transcription factor 4 (OCT4), SOX2, and NANOG gene expression. The fd-ECM induced chondrogenic differentiation in ad-MSCs compared to control ad-MSCs. Loss of function studies, through the use of siRNA and a mutant Notch1 construct, revealed that ECM-mediated ad-MSCs chondrogenesis requires Notch1 and β-catenin signaling. The fd-ECM also showed anti-senescence effects on ad-MSCs. The fd-ECM is a promising approach for inducing chondrogenesis in ad-MSCs and chondrogenic differentiated ad-MSCs could be used in stem cell therapy procedures. PMID:27527147

  11. The Extracellular Protease Matrix Metalloproteinase-9 Is Activated by Inhibitory Avoidance Learning and Required for Long-Term Memory

    ERIC Educational Resources Information Center

    Nagy, Vanja; Bozdagi, Ozlem; Huntley, George W.

    2007-01-01

    Matrix metalloproteinases (MMPs) are a family of extracellularly acting proteolytic enzymes with well-recognized roles in plasticity and remodeling of synaptic circuits during brain development and following brain injury. However, it is now becoming increasingly apparent that MMPs also function in normal, nonpathological synaptic plasticity of the…

  12. Varying RGD concentration and cell phenotype alters the expression of extracellular matrix genes in vocal fold fibroblasts.

    PubMed

    Kosinski, Aaron M; Sivasankar, M Preeti; Panitch, Alyssa

    2015-09-01

    The impact of RGD integrin binding-peptide concentration and cell phenotype on directing extracellular matrix (ECM) gene expression in vocal fold fibroblasts is little understood. Less is known about cell response to RGD concentration on a biomaterial when fibroblasts are in a scar-like environment compared to a healthy environment. We investigated the effects of varying RGD integrin-binding peptide surface concentration on ECM gene expression of elastin, collagen type 3 alpha 1, decorin, fibronectin, hyaluronan synthase 2, and collagen type 1 alpha 2 in scarred and unscarred immortalized human vocal fold fibroblasts (I-HVFFs). Phenotype and RGD concentration affected ECM gene expression. Phenotype change from healthy to myofibroblast-like resulted in ECM gene up-regulation for all genes tested, except for decorin. Systematically altering RGD concentration affected the expression of elastin and collagen type 3 alpha 1 in a myofibroblast phenotype. Specifically greater up-regulation in gene expression was observed with higher RGD concentrations. This research demonstrates that controlling RGD concentration may influence ECM gene expression levels in fibroblasts. Such knowledge is critical in developing the next generation of bioactive materials that, when implanted into sites of tissue damage and scarring, will direct cells to regenerate healthy tissues with normal ECM ratios and morphologies. PMID:25778824

  13. Extracellular matrix in the trabecular meshwork: Intraocular pressure regulation and dysregulation in glaucoma

    PubMed Central

    Vranka, Janice A.; Kelley, Mary J.; Acott, Ted S.; Keller, Kate E.

    2014-01-01

    The trabecular meshwork (TM) is located in the anterior segment of the eye and is responsible for regulating the outflow of aqueous humor. Increased resistance to aqueous outflow causes intraocular pressure to increase, which is the primary risk factor for glaucoma. TM cells reside on a series of fenestrated beams and sheets through which the aqueous humor flows to exit the anterior chamber via Schlemm’s canal. The outer trabecular cells are phagocytic and are thought to function as a pre-filter. However, most of the outflow resistance is thought to be from the extracellular matrix (ECM) of the juxtacanalicular region, the deepest portion of the TM, and from the inner wall basement membrane of Schlemm’s canal. It is becoming increasingly evident that the extracellular milieu is important in maintaining the integrity of the TM. Not only have ultrastructural changes been observed in the ECM of the TM in glaucoma, and a significant number of mutations in ECM genes are known to be associated with glaucoma, but the stiffness of glaucomatous TM appears to be greater than that of normal tissue. Additionally, TGFβ2 has been found to be elevated in the aqueous humor of glaucoma patients and is assumed to be involved in ECM changes deep with the juxtacanalicular region of the TM. This review summarizes the current literature on trabecular ECM as well as the development and function of the TM. Animal models and organ culture models targeting specific ECM molecules to investigate the mechanisms of glaucoma are described. Finally, the growing number of mutations that have been identified in ECM genes and genes that modulate ECM in humans with glaucoma are documented. PMID:25819459

  14. Activation of a 66-kilodalton human endothelial cell matrix metalloprotease by Streptococcus pyogenes extracellular cysteine protease.

    PubMed

    Burns, E H; Marciel, A M; Musser, J M

    1996-11-01

    Human umbilical vein endothelial cells (HUVECs) were used to gain insight into the molecular mechanism whereby the major extracellular protease from group A streptococci damages host tissue. HUVECs exposed to streptococcal cysteine protease (SCP) for various times exhibited cytopathic effect and cell detachment from the culture vessel. Gelatin substrate zymography showed that a time- and concentration-dependent increase in the level of activity of an approximately 66-kDa gelatinase occurred in culture medium taken from cells exposed to enzymatically active SCP. This gelatinase comigrated in gelatin zymograms with the activated form of purified recombinant matrix metalloprotease 2 (MMP-2) and had type IV collagenase activity. In contrast, medium taken from cells exposed to inactivated (boiled) SCP and cells exposed to SCP inhibited by treatment with N-benzyloxycarbonyl-leucyl-valyl-glycine diazomethyl ketone lacked the 66-kDa gelatinase. Appearance of the 66-kDa gelatinase activity was also prevented by 1,10-phenanthroline, a zinc chelator and MMP inhibitor. Inasmuch as proteolytically active SCP is required for the emergence of this gelatinase and MMP activation occurs by proteolytic processing, the 66-kDa gelatinase may be a proteolytic cleavage product of a latent MMP expressed extracellularly by HUVECs. Direct SCP treatment of culture supernatant taken from HUVECs not exposed to SCP also produced the 66-kDa gelatinase. The data show that SCP activates an MMP produced by human endothelial cells, a process that may contribute to endothelial cell damage, tissue destruction, and hemodynamic derangement observed in some patients with severe, invasive group A streptococcal infection. PMID:8890235

  15. Activation of a 66-kilodalton human endothelial cell matrix metalloprotease by Streptococcus pyogenes extracellular cysteine protease.

    PubMed Central

    Burns, E H; Marciel, A M; Musser, J M

    1996-01-01

    Human umbilical vein endothelial cells (HUVECs) were used to gain insight into the molecular mechanism whereby the major extracellular protease from group A streptococci damages host tissue. HUVECs exposed to streptococcal cysteine protease (SCP) for various times exhibited cytopathic effect and cell detachment from the culture vessel. Gelatin substrate zymography showed that a time- and concentration-dependent increase in the level of activity of an approximately 66-kDa gelatinase occurred in culture medium taken from cells exposed to enzymatically active SCP. This gelatinase comigrated in gelatin zymograms with the activated form of purified recombinant matrix metalloprotease 2 (MMP-2) and had type IV collagenase activity. In contrast, medium taken from cells exposed to inactivated (boiled) SCP and cells exposed to SCP inhibited by treatment with N-benzyloxycarbonyl-leucyl-valyl-glycine diazomethyl ketone lacked the 66-kDa gelatinase. Appearance of the 66-kDa gelatinase activity was also prevented by 1,10-phenanthroline, a zinc chelator and MMP inhibitor. Inasmuch as proteolytically active SCP is required for the emergence of this gelatinase and MMP activation occurs by proteolytic processing, the 66-kDa gelatinase may be a proteolytic cleavage product of a latent MMP expressed extracellularly by HUVECs. Direct SCP treatment of culture supernatant taken from HUVECs not exposed to SCP also produced the 66-kDa gelatinase. The data show that SCP activates an MMP produced by human endothelial cells, a process that may contribute to endothelial cell damage, tissue destruction, and hemodynamic derangement observed in some patients with severe, invasive group A streptococcal infection. PMID:8890235

  16. Changes in the Chondrocyte and Extracellular Matrix Proteome during Post-natal Mouse Cartilage Development*

    PubMed Central

    Wilson, Richard; Norris, Emma L.; Brachvogel, Bent; Angelucci, Constanza; Zivkovic, Snezana; Gordon, Lavinia; Bernardo, Bianca C.; Stermann, Jacek; Sekiguchi, Kiyotoshi; Gorman, Jeffrey J.; Bateman, John F.

    2012-01-01

    Skeletal growth by endochondral ossification involves tightly coordinated chondrocyte differentiation that creates reserve, proliferating, prehypertrophic, and hypertrophic cartilage zones in the growth plate. Many human skeletal disorders result from mutations in cartilage extracellular matrix (ECM) components that compromise both ECM architecture and chondrocyte function. Understanding normal cartilage development, composition, and structure is therefore vital to unravel these disease mechanisms. To study this intricate process in vivo by proteomics, we analyzed mouse femoral head cartilage at developmental stages enriched in either immature chondrocytes or maturing/hypertrophic chondrocytes (post-natal days 3 and 21, respectively). Using LTQ-Orbitrap tandem mass spectrometry, we identified 703 cartilage proteins. Differentially abundant proteins (q < 0.01) included prototypic markers for both early and late chondrocyte differentiation (epiphycan and collagen X, respectively) and novel ECM and cell adhesion proteins with no previously described roles in cartilage development (tenascin X, vitrin, Urb, emilin-1, and the sushi repeat-containing proteins SRPX and SRPX2). Meta-analysis of cartilage development in vivo and an in vitro chondrocyte culture model (Wilson, R., Diseberg, A. F., Gordon, L., Zivkovic, S., Tatarczuch, L., Mackie, E. J., Gorman, J. J., and Bateman, J. F. (2010) Comprehensive profiling of cartilage extracellular matrix formation and maturation using sequential extraction and label-free quantitative proteomics. Mol. Cell. Proteomics 9, 1296–1313) identified components involved in both systems, such as Urb, and components with specific roles in vivo, including vitrin and CILP-2 (cartilage intermediate layer protein-2). Immunolocalization of Urb, vitrin, and CILP-2 indicated specific roles at different maturation stages. In addition to ECM-related changes, we provide the first biochemical evidence of changing endoplasmic reticulum function during

  17. A Gene Expression-Based Comparison of Cell Adhesion to Extracellular Matrix and RGD-Terminated Monolayers

    PubMed Central

    Sobers, Courtney J.; Wood, Sarah E.; Mrksich, Milan

    2015-01-01

    This work uses global gene expression analysis to compare the extent to which model substrates presenting peptide adhesion motifs mimic the use of conventional extracellular matrix protein coated substrates for cell culture. We compared the transcriptional activities of genes in cells that were cultured on matrix-coated substrates with those cultured on self-assembled monolayers presenting either a linear or cyclic RGD peptide. Cells adherent to cyclic RGD were most similar to those cultured on native ECM, while cells cultured on monolayers presenting the linear RGD peptide had transcriptional activities that were more similar to cells cultured on the uncoated substrates. This study suggests that biomaterials presenting the cyclic RGD peptide are substantially better mimics of extracellular matrix than are uncoated materials or materials presenting the common linear RGD peptide. PMID:25818445

  18. Sec24D-Dependent Transport of Extracellular Matrix Proteins Is Required for Zebrafish Skeletal Morphogenesis

    PubMed Central

    Sarmah, Swapnalee; Barrallo-Gimeno, Alejandro; Melville, David B.; Topczewski, Jacek; Solnica-Krezel, Lilianna; Knapik, Ela W.

    2010-01-01

    Protein transport from endoplasmic reticulum (ER) to Golgi is primarily conducted by coated vesicular carriers such as COPII. Here, we describe zebrafish bulldog mutations that disrupt the function of the cargo adaptor Sec24D, an integral component of the COPII complex. We show that Sec24D is essential for secretion of cartilage matrix proteins, whereas the preceding development of craniofacial primordia and pre-chondrogenic condensations does not depend on this isoform. Bulldog chondrocytes fail to secrete type II collagen and matrilin to extracellular matrix (ECM), but membrane bound receptor β1-Integrin and Cadherins appear to leave ER in Sec24D-independent fashion. Consequently, Sec24D-deficient cells accumulate proteins in the distended ER, although a subset of ER compartments and Golgi complexes as visualized by electron microscopy and NBD C6-ceramide staining appear functional. Consistent with the backlog of proteins in the ER, chondrocytes activate the ER stress response machinery and significantly upregulate BiP transcription. Failure of ECM secretion hinders chondroblast intercalations thus resulting in small and malformed cartilages and severe craniofacial dysmorphology. This defect is specific to Sec24D mutants since knockdown of Sec24C, a close paralog of Sec24D, does not result in craniofacial cartilage dysmorphology. However, craniofacial development in double Sec24C/Sec24D-deficient animals is arrested earlier than in bulldog/sec24d, suggesting that Sec24C can compensate for loss of Sec24D at initial stages of chondrogenesis, but Sec24D is indispensable for chondrocyte maturation. Our study presents the first developmental perspective on Sec24D function and establishes Sec24D as a strong candidate for cartilage maintenance diseases and craniofacial birth defects. PMID:20442775

  19. Cardiac differentiation of cardiosphere-derived cells in scaffolds mimicking morphology of the cardiac extracellular matrix.

    PubMed

    Xu, Yanyi; Patnaik, Sourav; Guo, Xiaolei; Li, Zhenqing; Lo, Wilson; Butler, Ryan; Claude, Andrew; Liu, Zhenguo; Zhang, Ge; Liao, Jun; Anderson, Peter M; Guan, Jianjun

    2014-08-01

    Stem cell therapy has the potential to regenerate heart tissue after myocardial infarction (MI). The regeneration is dependent upon cardiac differentiation of the delivered stem cells. We hypothesized that timing of the stem cell delivery determines the extent of cardiac differentiation as cell differentiation is dependent on matrix properties such as biomechanics, structure and morphology, and these properties in cardiac extracellular matrix (ECM) continuously vary with time after MI. In order to elucidate the relationship between ECM properties and cardiac differentiation, we created an in vitro model based on ECM-mimicking fibers and a type of cardiac progenitor cell, cardiosphere-derived cells (CDCs). A simultaneous fiber electrospinning and cell electrospraying technique was utilized to fabricate constructs. By blending a highly soft hydrogel with a relatively stiff polyurethane and modulating fabrication parameters, tissue constructs with similar cell adhesion property but different global modulus, single fiber modulus, fiber density and fiber alignment were achieved. The CDCs remained alive within the constructs during a 1week culture period. CDC cardiac differentiation was dependent on the scaffold modulus, fiber volume fraction and fiber alignment. Two constructs with relatively low scaffold modulus, ∼50-60kPa, most significantly directed the CDC differentiation into mature cardiomyocytes as evidenced by gene expressions of cardiac troponin T (cTnT), calcium channel (CACNA1c) and cardiac myosin heavy chain (MYH6), and protein expressions of cardiac troponin I (cTnI) and connexin 43 (CX43). Of these two low-modulus constructs, the extent of differentiation was greater for lower fiber alignment and higher fiber volume fraction. These results suggest that cardiac ECM properties may have an effect on cardiac differentiation of delivered stem cells. PMID:24769114

  20. Bone Regeneration Using Hydroxyapatite Sponge Scaffolds with In Vivo Deposited Extracellular Matrix.

    PubMed

    Ventura, Reiza Dolendo; Padalhin, Andrew Reyes; Min, Young-Ki; Lee, Byong-Taek

    2015-11-01

    There is currently an increased interest in studying the extracellular matrix (ECM) and its potential applications for tissue engineering and regenerative medicine. The ECM plays an important role by providing adhesive substrates to cells during migration, morphogenesis, differentiation, and homeostasis by signaling biochemical and biomechanical cues to cells. In this study, the ECM was incorporated into hydroxyapatite by implanting sponge replica scaffolds in subcutaneous pockets in rats, and the implants were tested for bone regeneration potential. The resulting scaffolds were characterized using scanning electron microscopy, confocal microscopy, DNA and RNA quantification, tissue staining, energy dispersive X-ray spectroscopy analysis, compressive strength testing, porosity, and pore size distribution analysis using bare scaffolds as a control reference. Biocompatibility was assessed using MC3T3-E1 preosteoblast cells and in vivo studies were carried out by implanting decellularized scaffolds in 11 mm radial defects in New Zealand rabbits for 4 and 8 weeks to determine the effect of the in vivo deposited ECM. Material characterization indicated that a 2-week decellularized scaffold was the best among the samples, with an evenly distributed ECM visible on hematoxylin and eosin-stained tissue sections, a compressive strength of 2.53 ± 0.68 MPa, a porosity of 58.08 ± 3.32% and a pore size distribution range of 10-150 μm. In vivo results showed no severe inflammation, with increased cell infiltration followed by dense matrix deposition after 4 weeks and new bone formation at 8 weeks. The results indicate that incorporation of an in vivo deposited ECM into ceramic scaffolds can potentially improve bone regeneration. PMID:26228909

  1. Hypoxia and Extracellular Matrix Proteins Influence Angiogenesis and Lymphangiogenesis in Mouse Embryoid Bodies

    PubMed Central

    Foskett, Andrea M.; Ezekiel, Uthayashanker R.; Trzeciakowski, Jerome P.; Zawieja, David C.; Muthuchamy, Mariappan

    2011-01-01

    Regulatory mechanisms for angiogenesis are relatively well established compared to lymphangiogenesis. Few studies have shown that a combination of vascular endothelial growth factor VEGF-A/C with hypoxia or collagen matrix promotes lymphatic structures along with blood vessel development in mouse embryoid bodies (EB). In this study we tested the hypothesis that while hypoxia combined with prolonged VEGF-A/C treatment would induce early lymphangiogenesis in addition to angiogenesis in mouse EBs, under similar conditions specific extracellular matrix (ECM) proteins would promote lymphatic vessel-like structures over angiogenesis. EBs were subjected to four conditions and were maintained under normoxia and hypoxia (21% and 2.6% O2, respectively) with or without VEGF-A/C. Microarray analyses of normoxic and hypoxic EBs, and immunofluorescence data showed very low expression of early lymphatic endothelial cell (LEC) markers, lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1), and prospero-related homeobox 1 (Prox1) at early time points. Double immunofluorescence using MECA-32 and Prox1/LYVE1 demonstrated that combined hypoxia and VEGF-A/C treatment promoted formation of blood vessel-like structures, whereas only Prox1+/LYVE1+ LECs were detected in EBs at E22.5. Furthermore, EBs were grown on laminin or collagen-I coated plates and were subjected to the four treatments as described above. Results revealed that LECs in EBs at E36.5 attached better to collagen-I, resulting in an organized network of lymphatic vessel-like structures as compared to EBs grown on laminin. However, blood vessel-like structures were less favored under these same conditions. Collectively, our data demonstrate that hypoxia combined with growth factors promotes angiogenesis, whereas combination of these conditions with specific ECM proteins favors lymphangiogenesis processes in mouse EBs. PMID:22194726

  2. How do fibroblasts translate mechanical signals into changes in extracellular matrix production?

    PubMed

    Chiquet, Matthias; Renedo, Ana Sarasa; Huber, François; Flück, Martin

    2003-03-01

    Mechanical forces are important regulators of connective tissue homeostasis. Our recent experiments in vivo indicate that externally applied mechanical load can lead to the rapid and sequential induction of distinct extracellular matrix (ECM) components in fibroblasts, rather than to a generalized hypertrophic response. Thus, ECM composition seems to be adapted specifically to changes in load. Mechanical stress can regulate the production of ECM proteins indirectly, by stimulating the release of a paracrine growth factor, or directly, by triggering an intracellular signalling pathway that activates the gene. We have evidence that tenascin-C is an ECM component directly regulated by mechanical stress: induction of its mRNA in stretched fibroblasts is rapid both in vivo and in vitro, does not depend on prior protein synthesis, and is not mediated by factors released into the medium. Fibroblasts sense force-induced deformations (strains) in their ECM. Findings by other researchers indicate that integrins within cell-matrix adhesions can act as 'strain gauges', triggering MAPK and NF-kappaB pathways in response to changes in mechanical stress. Our results indicate that cytoskeletal 'pre-stress' is important for mechanotransduction to work: relaxation of the cytoskeleton (e.g. by inhibiting Rho-dependent kinase) suppresses induction of the tenascin-C gene by cyclic stretch, and hence desensitizes the fibroblasts to mechanical signals. On the level of the ECM genes, we identified related enhancer sequences that respond to static stretch in both the tenascin-C and the collagen XII promoter. In the case of the tenascin-C gene, different promoter elements might be involved in induction by cyclic stretch. Thus, different mechanical signals seem to regulate distinct ECM genes in complex ways. PMID:12714044

  3. Extracellular Matrix Invasion in Metastases and Angiogenesis: Commentary on the Matrigel "Chemoinvasion Assay".

    PubMed

    Albini, Adriana

    2016-08-15

    Invasive and metastatic cells must cross the basement membrane's extracellular matrix to disseminate to