WorldWideScience

Sample records for human fibrinous matrix

  1. Human Kunitz-type protease inhibitor engineered for enhanced matrix retention extends longevity of fibrin biomaterials.

    Science.gov (United States)

    Briquez, Priscilla S; Lorentz, Kristen M; Larsson, Hans M; Frey, Peter; Hubbell, Jeffrey A

    2017-08-01

    Aprotinin is a broad-spectrum serine protease inhibitor used in the clinic as an anti-fibrinolytic agent in fibrin-based tissue sealants. However, upon re-exposure, some patients suffer from hypersensitivity immune reactions likely related to the bovine origin of aprotinin. Here, we aimed to develop a human-derived substitute to aprotinin. Based on sequence homology analyses, we identified the Kunitz-type protease inhibitor (KPI) domain of human amyloid-β A4 precursor protein as being a potential candidate. While KPI has a lower intrinsic anti-fibrinolytic activity than aprotinin, we reasoned that its efficacy is additionally limited by its fast release from fibrin material, just as aprotinin's is. Thus, we engineered KPI variants for controlled retention in fibrin biomaterials, using either covalent binding through incorporation of a substrate for the coagulation transglutaminase Factor XIIIa or through engineering of extracellular matrix protein super-affinity domains for sequestration into fibrin. We showed that both engineered KPI variants significantly slowed plasmin-mediated fibrinolysis in vitro, outperforming aprotinin. In vivo, our best engineered KPI variant (incorporating the transglutaminase substrate) extended fibrin matrix longevity by 50%, at a dose at which aprotinin did not show efficacy, thus qualifying it as a competitive substitute of aprotinin in fibrin sealants. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  2. Complex interactions between human myoblasts and the surrounding 3D fibrin-based matrix.

    Directory of Open Access Journals (Sweden)

    Stéphane Chiron

    Full Text Available Anchorage of muscle cells to the extracellular matrix is crucial for a range of fundamental biological processes including migration, survival and differentiation. Three-dimensional (3D culture has been proposed to provide a more physiological in vitro model of muscle growth and differentiation than routine 2D cultures. However, muscle cell adhesion and cell-matrix interplay of engineered muscle tissue remain to be determined. We have characterized cell-matrix interactions in 3D muscle culture and analyzed their consequences on cell differentiation. Human myoblasts were embedded in a fibrin matrix cast between two posts, cultured until confluence, and then induced to differentiate. Myoblasts in 3D aligned along the longitudinal axis of the gel. They displayed actin stress fibers evenly distributed around the nucleus and a cortical mesh of thin actin filaments. Adhesion sites in 3D were smaller in size than in rigid 2D culture but expression of adhesion site proteins, including α5 integrin and vinculin, was higher in 3D compared with 2D (p<0.05. Myoblasts and myotubes in 3D exhibited thicker and ellipsoid nuclei instead of the thin disk-like shape of the nuclei in 2D (p<0.001. Differentiation kinetics were faster in 3D as demonstrated by higher mRNA concentrations of α-actinin and myosin. More important, the elastic modulus of engineered muscle tissues increased significantly from 3.5 ± 0.8 to 7.4 ± 4.7 kPa during proliferation (p<0.05 and reached 12.2 ± 6.0 kPa during differentiation (p<0.05, thus attesting the increase of matrix stiffness during proliferation and differentiation of the myocytes. In conclusion, we reported modulations of the adhesion complexes, the actin cytoskeleton and nuclear shape in 3D compared with routine 2D muscle culture. These findings point to complex interactions between muscle cells and the surrounding matrix with dynamic regulation of the cell-matrix stiffness.

  3. Urokinase receptor expression on human microvascular endothelial cells is increased by hypoxia: Implications for capillary-like tube formation in a fibrin matrix

    NARCIS (Netherlands)

    Kroon, M.E.; Koolwijk, P.; Vecht, B. van der; Hinsbergh, V.W.M. van

    2000-01-01

    Hypoxia stimulates angiogenesis, the formation of new blood vessels. This study evaluates the direct effect of hypoxia (1% oxygen) on the angiogenic response of human microvascular endothelial cells (hMVECs) seeded on top of a 3-dimensional fibrin matrix, hMVECs stimulated with fibroblast growth

  4. Reconstruction of living bilayer human skin equivalent utilizing human fibrin as a scaffold.

    Science.gov (United States)

    Mazlyzam, A L; Aminuddin, B S; Fuzina, N H; Norhayati, M M; Fauziah, O; Isa, M R; Saim, L; Ruszymah, B H I

    2007-05-01

    Our aim of this study was to develop a new methodology for constructing a bilayer human skin equivalent to create a more clinical compliance skin graft composite for the treatment of various skin defects. We utilized human plasma derived fibrin as the scaffold for the development of a living bilayer human skin equivalent: fibrin-fibroblast and fibrin-keratinocyte (B-FF/FK SE). Skin cells from six consented patients were culture-expanded to passage 1. For B-FF/FK SE formation, human fibroblasts were embedded in human fibrin matrix and subsequently another layer of human keratinocytes in human fibrin matrix was stacked on top. The B-FF/FK SE was then transplanted to athymic mice model for 4 weeks to evaluate its regeneration and clinical performance. The in vivo B-FF/FK SE has similar properties as native human skin by histological analysis and expression of basal Keratin 14 gene in the epidermal layer and Collagen type I gene in the dermal layer. Electron microscopy analysis of in vivo B-FF/FK SE showed well-formed and continuous epidermal-dermal junction. We have successfully developed a technique to engineer living bilayer human skin equivalent using human fibrin matrix. The utilization of culture-expanded human skin cells and fibrin matrix from human blood will allow a fully autologous human skin equivalent construction.

  5. Platelet-rich fibrin matrix for facial plastic surgery.

    Science.gov (United States)

    Sclafani, Anthony P; Saman, Masoud

    2012-05-01

    Platelets are known primarily for their role in hemostasis, but there is increasing interest in the effect of platelets on wound healing. Platelet isolates such as platelet-rich plasma have been advocated to enhance and accelerate wound healing. This article describes the use of a novel preparation, platelet-rich fibrin matrix (PRFM), for facial plastic surgery applications such as volume augmentation, fat transfer supplementation, and as an adjunct to open surgical procedures. Copyright © 2012 Elsevier Inc. All rights reserved.

  6. Fibronectin alters the rate of formation and structure of the fibrin matrix.

    Science.gov (United States)

    Ramanathan, Anand; Karuri, Nancy

    2014-01-10

    Plasma fibronectin is a vital component of the fibrin clot; however its role on clot structure is not clearly understood. The goal of this study was to examine the influence of fibronectin on the kinetics of formation, structural characteristics and composition of reconstituted fibrin clots or fibrin matrices. Fibrin matrices were formed by adding thrombin to 1, 2 or 4 mg/ml fibrinogen supplemented with 0-0.4 mg/ml fibronectin. The rate of fibrin matrix formation was then monitored by measuring light absorbance properties at different time points. Confocal microscopy of fluorescein conjugated fibrinogen was used to visualize the structural characteristics of fibrin matrices. The amount of fibronectin in fibrin matrices was determined through electrophoresis and immunoblotting of solubilized matrices. Fibronectin concentration positively correlated with the initial rate of fibrin matrix formation and with steady state light absorbance values of fibrin matrices. An increase in fibronectin concentration resulted in thinner and denser fibers in the fibrin matrices. Electrophoresis and immunoblotting showed that fibronectin was covalently and non-covalently bound to fibrin matrices and in the form of high molecular weight multimers. The formation of fibronectin multimers was attributed to cross-linking of fibronectin by trace amounts Factor XIIIa. These findings are novel because they link results from light absorbance studies to microcopy analyses and demonstrate an influence of fibronectin on fibrin matrix structural characteristics. This data is important in developing therapies that destabilize fibrin clots. Copyright © 2014. Published by Elsevier Inc.

  7. [Conservative anal fistula treatment with collagenic plug and human fibrin sealant. Preliminary results].

    Science.gov (United States)

    Gubitosi, A; Moccia, G; Malinconico, F A; Docimo, G; Ruggiero, R; Iside, G; Avenia, N; Docimo, L; Foroni, F; Gilio, F; Sparavigna, L; Agresti, M

    2009-01-01

    The authors, on the basis of a long clinical experience with human fibrin glue in general surgery, compared two different extracellular matrix (collagen), Surgisis and TissueDura, with human fibrin glue, applied during the operation, and sometimes in postoperative, to obtain the healing of perianal fistulas. The collagenic extracellular matrix provides, according to the rationale suggested, an optimal three-dimensional structure for the fibroblastic implant and neoangiogenesis, hence for the fistula "fibrotizzation" and closure. The encouraging results for transphincteric fistulas and a simple and easy technique push to researchers on samples statistically significant.

  8. A recently developed bifacial platelet-rich fibrin matrix

    Directory of Open Access Journals (Sweden)

    E Lucarelli

    2010-07-01

    Full Text Available Platelet-rich plasma (PRP is used clinically in liquid or gel form to promote tissue repair. Because of the poor mechanical properties, conventional PRP is often difficult to handle when used in clinical settings and requires secure implantation in a specific site, otherwise when released growth factors could be washed out during an operation. In this study, we analyzed the end product of a recently developed commercially available system (FIBRINET®, which is a dense pliable, platelet-rich fibrin matrix (PRFM. We characterized the mechanical properties of PRFM and tested whether PRFM releases growth factors and whether released factors induce the proliferation of mesenchymal stem cells (MSC. Mechanical properties as well as platelet distribution were evaluated in PRFM. PRFM demonstrated robust mechanical properties, with a tear elastic modulus of 937.3 + 314.6 kPa, stress at a break of 1476.0 + 526.3 kPa, and an elongation at break of 146.3 + 33.8 %. PRFM maintained its mechanical properties throughout the testing process. Microscopic observations showed that the platelets were localized on one side of the matrix. Elevated levels of PDGF-AA, PDGF-AB, EGF, VEGF, bFGF and TGF-β1 were measured in the day 1-conditioned media (CM of PRFM and growth factor levels decreased thereafter. BMP2 and BMP7 were not detectable. MSC culture media supplemented with 20% PRFM-CM stimulated MSC cell proliferation; at 24 and 48 hours the induction of the proliferation was significantly greater than the induction obtained with media supplemented with 20% foetal bovine serum. The present study shows that the production of a dense, physically robust PRFM made through high-speed centrifugation of intact platelets and fibrin in the absence of exogenous thrombin yields a potential tool for accelerating tissue repair.

  9. Platelet-rich fibrin matrix improves wound angiogenesis via inducing endothelial cell proliferation.

    Science.gov (United States)

    Roy, Sashwati; Driggs, Jason; Elgharably, Haytham; Biswas, Sabyasachi; Findley, Muna; Khanna, Savita; Gnyawali, Urmila; Bergdall, Valerie K; Sen, Chandan K

    2011-11-01

    The economic, social, and public health burden of chronic ulcers and other compromised wounds is enormous and rapidly increasing with the aging population. The growth factors derived from platelets play an important role in tissue remodeling including neovascularization. Platelet-rich plasma (PRP) has been utilized and studied for the last four decades. Platelet gel and fibrin sealant, derived from PRP mixed with thrombin and calcium chloride, have been exogenously applied to tissues to promote wound healing, bone growth, hemostasis, and tissue sealing. In this study, we first characterized recovery and viability of as well as growth factor release from platelets in a novel preparation of platelet gel and fibrin matrix, namely platelet-rich fibrin matrix (PRFM). Next, the effect of PRFM application in a delayed model of ischemic wound angiogenesis was investigated. The study, for the first time, shows the kinetics of the viability of platelet-embedded fibrin matrix. A slow and steady release of growth factors from PRFM was observed. The vascular endothelial growth factor released from PRFM was primarily responsible for endothelial mitogenic response via extracellular signal-regulated protein kinase activation pathway. Finally, this preparation of PRFM effectively induced endothelial cell proliferation and improved wound angiogenesis in chronic wounds, providing evidence of probable mechanisms of action of PRFM in healing of chronic ulcers. 2011 by the Wound Healing Society.

  10. Cyclic compression maintains viability and induces chondrogenesis of human mesenchymal stem cells in fibrin gel scaffolds.

    Science.gov (United States)

    Pelaez, Daniel; Huang, Chun-Yuh Charles; Cheung, Herman S

    2009-01-01

    Mechanical loading has long been shown to modulate cartilage-specific extracellular matrix synthesis. With joint motion, cartilage can experience mechanical loading in the form of compressive, tensile or shearing load, and hydrostatic pressure. Recent studies have demonstrated the capacity of unconfined cyclic compression to induce chondrogenic differentiation of human mesenchymal stem cell (hMSC) in agarose culture. However, the use of a nonbiodegradable material such as agarose limits the applicability of these constructs. Of the possible biocompatible materials available for tissue engineering, fibrin is a natural regenerative scaffold, which possesses several desired characteristics including a controllable degradation rate and low immunogenicity. The objective of the present study was to determine the capability of fibrin gels for supporting chondrogenesis of hMSCs under cyclic compression. To optimize the system, three concentrations of fibrin gel (40, 60, and 80 mg/mL) and three different stimulus frequencies (0.1, 0.5, and 1.0 Hz) were used to examine the effects of cyclic compression on viability, proliferation and chondrogenic differentiation of hMSCs. Our results show that cyclic compression (10% strain) at frequencies >0.5 Hz and gel concentration of 40 mg/mL fibrinogen appears to maintain cellular viability within scaffolds. Similarly, variations in gel component concentration and stimulus frequency can be modified such that a significant chondrogenic response can be achieved by hMSC in fibrin constructs after 8 h of compression spread out over 2 days. This study demonstrates the suitability of fibrin gel for supporting the cyclic compression-induced chondrogenesis of mesenchymal stem cells.

  11. Neutrophil Extracellular Traps and Fibrin in Otitis Media: Analysis of Human and Chinchilla Temporal Bones.

    Science.gov (United States)

    Schachern, Patricia A; Kwon, Geeyoun; Briles, David E; Ferrieri, Patricia; Juhn, Steven; Cureoglu, Sebahattin; Paparella, Michael M; Tsuprun, Vladimir

    2017-10-01

    Bacterial resistance in acute otitis can result in bacterial persistence and biofilm formation, triggering chronic and recurrent infections. To investigate the middle ear inflammatory response to bacterial infection in human and chinchilla temporal bones. Six chinchillas underwent intrabullar inoculations with 0.5 mL of 106 colony-forming units (CFUs) of Streptococcus pneumoniae, serotype 2. Two days later, we counted bacteria in middle ear effusions postmortem. One ear from each chinchilla was processed in paraffin and sectioned at 5 µm. The opposite ear was embedded in epoxy resin, sectioned at a thickness of 1 µm, and stained with toluidine blue. In addition, we examined human temporal bones from 2 deceased donors with clinical histories of otitis media (1 with acute onset otitis media, 1 with recurrent infection). Temporal bones had been previously removed at autopsy, processed, embedded in celloidin, and cut at a thickness of 20 µm. Sections of temporal bones from both chinchillas and humans were stained with hematoxylin-eosin and immunolabeled with antifibrin and antihistone H4 antibodies. Histopatological and imminohistochemical changes owing to otitis media. Bacterial counts in chinchilla middle ear effusions 2 days after inoculation were approximately 2 logs above initial inoculum counts. Both human and chinchilla middle ear effusions contained bacteria embedded in a fibrous matrix. Some fibers in the matrix showed positive staining with antifibrin antibody, others with antihistone H4 antibody. In acute and recurrent otitis media, fibrin and neutrophil extracellular traps (NETs) are part of the host inflammatory response to bacterial infection. In the early stages of otitis media the host defense system uses fibrin to entrap bacteria, and NETs function to eliminate bacteria. In chronic otitis media, fibrin and NETs appear to persist.

  12. Does tranexamic acid stabilised fibrin support the osteogenic differentiation of human periosteum derived cells?

    Directory of Open Access Journals (Sweden)

    J Demol

    2011-03-01

    Full Text Available Fibrin sealants have long been used as carrier for osteogenic cells in bone regeneration. However, it has not been demonstrated whether fibrin’s role is limited to delivering cells to the bone defect or whether fibrin enhances osteogenesis. This study investigated fibrin’s influence on the behaviour of human periosteum-derived cells (hPDCs when cultured in vitro under osteogenic conditions in two-dimensional (fibrin substrate and three-dimensional (fibrin carrier environments. Tranexamic acid (TEA was used to reduce fibrin degradation after investigating its effect on hPDCs in monolayer culture on plastic.TEA did not affect proliferation nor calcium deposition of hPDCs under these conditions. Expression profiles of specific osteogenic markers were also maintained within the presence of TEA, apart from reduced alkaline phosphatase (ALP expression (day 14. Compared to plastic, proliferation was upregulated on 2D fibrin substrates with a 220% higher DNA content by day 21. Gene expression was also altered, with significantly (p<0.05 decreased Runx2 (day 7 and ALP (day 14 expression and increased collagen I expression (day 14 and 21. In contrast to plastic, mineralisation was absent on fibrin substrates. Inside fibrin carriers, hPDCs were uniformly distributed. Moderate cell growth and reduced osteogenic marker expression was observed inside fibrin carriers. After 2 weeks, increased cell death was present in the carrier’s centre. In conclusion, fibrin negatively influences osteogenic differentiation, compared to culture plastic, but enhanced proliferation (at least in 2D cultures for hPDCs cultured in osteogenic conditions. TEA maintained the integrity of fibrin-based constructs, with minor effects on the osteogenic differentiation of hPDCs.

  13. Fibrin promotes proliferation and matrix production of intervertebral disc cells cultured in three-dimensional poly(lactic-co-glycolic acid) scaffold.

    Science.gov (United States)

    Sha'ban, Munirah; Yoon, Sun Jung; Ko, Youn Kyung; Ha, Hyun Jung; Kim, Soon Hee; So, Jung Won; Idrus, Ruszymah Bt Hj; Khang, Gilson

    2008-01-01

    Previously, we have proven that fibrin and poly(lactic-co-glycolic acid) (PLGA) scaffolds facilitate cell proliferation, matrix production and early chondrogenesis of rabbit articular chondrocytes in in vitro and in vivo experiments. In this study, we evaluated the potential of fibrin/PLGA scaffold for intervertebral disc (IVD) tissue engineering using annulus fibrosus (AF) and nucleus pulposus (NP) cells in relation to potential clinical application. PLGA scaffolds were soaked in cells-fibrin suspension and polymerized by dropping thrombin-sodium chloride (CaCl(2)) solution. A PLGA-cell complex without fibrin was used as control. Higher cellular proliferation activity was observed in fibrin/PLGA-seeded AF and NP cells at each time point of 3, 7, 14 and 7 days using the MTT assay. After 3 weeks in vitro incubation, fibrin/PLGA exhibited a firmer gross morphology than PLGA groups. A significant cartilaginous tissue formation was observed in fibrin/PLGA, as proven by the development of cells cluster of various sizes and three-dimensional (3D) cartilaginous histoarchitecture and the presence of proteoglycan-rich matrix and glycosaminoglycan (GAG). The sGAG production measured by 1,9-dimethylmethylene blue (DMMB) assay revealed greater sGAG production in fibrin/PLGA than PLGA group. Immunohistochemical analyses showed expressions of collagen type II, aggrecan core protein and collagen type I genes throughout in vitro culture in both fibrin/PLGA and PLGA. In conclusion, fibrin promotes cell proliferation, stable in vitro tissue morphology, superior cartilaginous tissue formation and sGAG production of AF and NP cells cultured in PLGA scaffold. The 3D porous PLGA scaffold-cell complexes using fibrin can provide a vehicle for delivery of cells to regenerate tissue-engineered IVD tissue.

  14. HIF-2α Expression Regulates Sprout Formation into 3D Fibrin Matrices in Prolonged Hypoxia in Human Microvascular Endothelial Cells.

    Science.gov (United States)

    Nauta, Tessa D; Duyndam, Monique C A; Weijers, Ester M; van Hinsbergh, Victor M W; Koolwijk, Pieter

    2016-01-01

    During short-term hypoxia, Hypoxia Inducible Factors (particular their subunits HIF-1α and HIF-2α) regulate the expression of many genes including the potent angiogenesis stimulator VEGF. However, in some pathological conditions chronic hypoxia occurs and is accompanied by reduced angiogenesis. We investigated the effect of prolonged hypoxia on the proliferation and sprouting ability of human microvascular endothelial cells and the involvement of the HIFs and Dll4/Notch signaling. Human microvascular endothelial cells (hMVECs), cultured at 20% oxygen for 14 days and seeded on top of 3D fibrin matrices, formed sprouts when stimulated with VEGF-A/TNFα. In contrast, hMVECs precultured at 1% oxygen for 14 days were viable and proliferative, but did not form sprouts into fibrin upon VEGF-A/TNFα stimulation at 1% oxygen. Silencing of HIF-2α with si-RNA partially restored the inhibition of endothelial sprouting, whereas HIF-1α or HIF-3α by si-RNA had no effect. No involvement of Dll4/Notch pathway in the inhibitory effect on endothelial sprouting by prolonged hypoxia was found. In addition, hypoxia decreased the production of urokinase-type plasminogen activator (uPA), needed for migration and invasion, without a significant effect on its inhibitor PAI-1. This was independent of HIF-2α, as si-HIF-2α did not counteract uPA reduction. Prolonged culturing of hMVECs at 1% oxygen inhibited endothelial sprouting into fibrin. Two independent mechanisms contribute. Silencing of HIF-2α with si-RNA partially restored the inhibition of endothelial sprouting pointing to a HIF-2α-dependent mechanism. In addition, reduction of uPA contributed to reduced endothelial tube formation in a fibrin matrix during prolonged hypoxia.

  15. Fabrication and physical and biological properties of fibrin gel derived from human plasma

    Science.gov (United States)

    Zhao, Haiguang; Ma, Lie; Zhou, Jie; Mao, Zhengwei; Gao, Changyou; Shen, Jiacong

    2008-03-01

    The fast development of tissue engineering and regenerative medicine drives the old biomaterials, for example, fibrin glue, to find new applications in these areas. Aiming at developing a commercially available hydrogel for cell entrapment and delivery, in this study we optimized the fabrication and gelation conditions of fibrin gel. Fibrinogen was isolated from human plasma by a freeze-thaw circle. Gelation of the fibrinogen was accomplished by mixing with thrombin. Absorbance of the fibrinogen/thrombin mixture at 550 nm as a function of reaction time was monitored by UV-VIS spectroscopy. It was found that the clotting time is significantly influenced by the thrombin concentration and the temperature, while less influenced by the fibrinogen concentration. After freeze-drying, the fibrin gel was characterized by scanning electron microscopy (SEM), revealing fibrous microstructure. Thermal gravimetric analysis found that the degradation temperature of the crosslinked fibrin gel starts from 288 °C, which is about 30 °C higher than that of the fibrinogen. The hydrogel has an initial water-uptake ratio of ~50, decreased to 30-40 after incubation in water for 11 h depending on the thrombin concentration. The fibrin gels lost their weights in PBS very rapidly, while slowly in DMEM/fetal bovine serum and DMEM. In vitro cell culture found that human fibroblasts could normally proliferate in the fibrin gel with spreading morphology. In conclusion, the fibrin gel containing higher concentration of fibrinogen (20 mg ml-1) and thrombin (5 U ml-1) has suitable gelation time and handling properties, and thus is applicable as a delivery vehicle for cells such as fibroblasts.

  16. In vivo chicken model for peripheral intravascular human fibrin clot detection

    International Nuclear Information System (INIS)

    Hui, K.Y.

    1988-01-01

    A chicken model was prepared that provides a simple and economical method of evaluating the use of fibrin-specific monoclonal antibody 64C5 in the detection of peripheral vascular thrombi. Human fibrin was clotted in segments of a chicken's femoral artery and vein prior to intravenous injection of radioiodinated antibody 64C5. After a 3-hr perfusion time, the thrombosed and contralateral control segments of the vessels were excised and counted for radioactivity. The radiolabeled 64C5 uptake ratio of the thrombosed segment to the control segment was 5.4 +/- 1.2 (p less than 0.007) in the femoral artery, and 3.8 +/- 1.1 (p less than 0.02) in the femoral vein. This in vivo chicken model may also find application in studies of targeting agents for human fibrin

  17. Safety and feasibility of platelet rich fibrin matrix injections for treatment of common urologic conditions

    Directory of Open Access Journals (Sweden)

    Ethan L Matz

    2018-01-01

    Full Text Available Purpose: Autologous platelet rich plasma (PRP is used increasingly in a variety of settings. PRP injections have been used for decades to improve angiogenesis and wound healing. They have also been offered commercially in urology with little to no data on safety or efficacy. PRP could theoretically improve multiple urologic conditions, such as erectile dysfunction (ED, Peyronie's disease (PD, and stress urinary incontinence (SUI. A concern with PRP, however, is early washout, a situation potentially avoided by conversion to platelet rich fibrin matrix (PRFM. Before clinical trials can be performed, safety analysis is desirable. We reviewed an initial series of patients receiving PRFM for urologic pathology to assess safety and feasibility. Materials and Methods: Data were reviewed for patients treated with PRFM at our center from November 2012 to July 2017. Patients were observed immediately post-injection and at follow-up for complications and tolerability. Where applicable, International Index of Erectile Function (IIEF-5 scores were reviewed before and after injections for ED and/or PD. Pad use data was collected pre/post injection for SUI. Results: Seventeen patients were identified, with a mean receipt of 2.1 injections per patient. Post-procedural minor adverse events were seen in 3 men, consisting of mild pain at injection site and mild penile bruising. No patients experienced complications at follow-up. No decline was observed in men completing pre/post IIEF-5 evaluations. Conclusions: PRFM appears to be a safe and feasible treatment modality in patients with urologic disease. Further placebo-controlled trials are warranted.

  18. Radiolabelled anti-human fibrin antibody: a new thrombus-detecting agent

    International Nuclear Information System (INIS)

    Bosnjakovic, V.; Jankovic, B.D.; Horvat, J.; Cvoric, J.

    1977-01-01

    Rabbit anti-human fibrin globulin (A.F.G.) was labelled with iodine ( 131 I) and used as a thrombus-detecting agent. 131 I-A.F.G. labelled thrombi were displayed by means of a gamma scintillation camera. Normal subjects and patients with thrombo-phlebitis of legs, acute fibrin depositions other than thrombi, and chronic varicosities were examined. The 131 I-A.F.G. technique detected both formed thrombi and those that were forming and could discriminate between acute thrombosis and chronic varicosities. Thrombo-phlebitis and extravascular fibrin depositions were best demonstrated between 24 and 72 hours after 131 I-A.F.G. injection. Radiolabelled A.F.G. in normal veins and chronic varicosities was best displayed within 6 hours of injection. (author)

  19. Quality Assessment of Platelet-Rich Fibrin-Like Matrix Prepared from Whole Blood Samples after Extended Storage

    Directory of Open Access Journals (Sweden)

    Hideo Kawabata

    2017-09-01

    Full Text Available The platelet-rich fibrin–like matrix (PRFM is usually prepared onsite and immediately used for regenerative therapy. Nonetheless, to meet the clinical necessity of preserving the PRFM without quality deterioration, we developed a method for preparation of PRFMs from short-term-stored whole blood (WB samples. In this study, to evaluate the practical expiration date of storage, we extended the storage time of WB samples from 2 to 7 days and assessed the quality of the resulting PRFMs. WB samples collected with acid-citrate-dextrose were stored with gentle agitation at ambient temperature. To prepare PRFMs, the stored WB samples were mixed with CaCl2 in glass tubes and centrifuged. Fibrin fiber networks, CD41 and CD62P expression, and Platelet Derived Growth Factor-BB (PDGF-BB levels were examined by scanning electron microscopy (SEM, flow cytometry, and an Enzyme-Linked ImmunoSorbent Assay (ELISA, respectively. Long-term storage had no significant effect on either blood cell counts or platelet functions tested. The resulting PRFMs were visually identical to freshly prepared ones. PDGF-BB levels did not markedly decrease in a time-dependent manner. However, fibrin fibers gradually became thinner after storage. Although the coagulation activity may diminish, we propose that PRFMs can be prepared—without evident loss of quality—from WB samples stored for up to 7 days by our previously developed method.

  20. Hematopoietic stem cell cytokines and fibroblast growth factor-2 stimulate human endothelial cell-pericyte tube co-assembly in 3D fibrin matrices under serum-free defined conditions.

    Directory of Open Access Journals (Sweden)

    Annie O Smith

    Full Text Available We describe a novel 3D fibrin matrix model using recombinant hematopoietic stem cell cytokines under serum-free defined conditions which promotes the assembly of human endothelial cell (EC tubes with co-associated pericytes. Individual ECs and pericytes are randomly mixed together and EC tubes form that is accompanied by pericyte recruitment to the EC tube abluminal surface over a 3-5 day period. These morphogenic processes are stimulated by a combination of the hematopoietic stem cell cytokines, stem cell factor, interleukin-3, stromal derived factor-1α, and Flt-3 ligand which are added in conjunction with fibroblast growth factor (FGF-2 into the fibrin matrix. In contrast, this tube morphogenic response does not occur under serum-free defined conditions when VEGF and FGF-2 are added together in the fibrin matrices. We recently demonstrated that VEGF and FGF-2 are able to prime EC tube morphogenic responses (i.e. added overnight prior to the morphogenic assay to hematopoietic stem cell cytokines in collagen matrices and, interestingly, they also prime EC tube morphogenesis in 3D fibrin matrices. EC-pericyte interactions in 3D fibrin matrices leads to marked vascular basement membrane assembly as demonstrated using immunofluorescence and transmission electron microscopy. Furthermore, we show that hematopoietic stem cell cytokines and pericytes stimulate EC sprouting in fibrin matrices in a manner dependent on the α5β1 integrin. This novel co-culture system, under serum-free defined conditions, allows for a molecular analysis of EC tube assembly, pericyte recruitment and maturation events in a critical ECM environment (i.e. fibrin matrices that regulates angiogenic events in postnatal life.

  1. Efficacy of platelet-rich fibrin matrix on viability of diced cartilage grafts in a rabbit model.

    Science.gov (United States)

    Güler, İsmail; Billur, Deniz; Aydin, Sevim; Kocatürk, Sinan

    2015-03-01

    The objective of this study was to compare the viability of cartilage grafts embedded in platelet-rich fibrin matrix (PRFM) wrapped with no material (bare diced cartilage grafts), oxidized methylcellulose (Surgicel), or acellular dermal tissue (AlloDerm). Experimental study. In this study, six New Zealand rabbits were used. Cartilage grafts including perichondrium were excised from each ear and diced into 2-mm-by 2-mm pieces. There were four comparison groups: 1) group A, diced cartilage (not wrapped with any material); 2) group B, diced cartilage wrapped with AlloDerm; 3) group C, diced cartilage grafts wrapped with Surgicel; and 4) group D, diced cartilage wrapped with PRFM. Four cartilage grafts were implanted under the skin at the back of each rabbit. All rabbits were sacrificed at the end of 10 weeks. The cartilages were stained with hematoxylin-eosin, Masson's Trichrome, and Orcein. After that, they were evaluated for the viability of chondrocytes, collagen content, fibrillar structure of matrix, and changes in peripheral tissues. When the viability of chondrocytes, the content of fiber in matrix, and changes in peripheral tissues were compared, the cartilage embedded in the PRFM group was statistically significantly higher than in the other groups (P < 0.05). We concluded that PRFM has significant advantages in ensuring the chondrocyte viability of diced cartilage grafts. It is also biocompatible, with relatively lesser inflammation and fibrosis. © 2014 The American Laryngological, Rhinological and Otological Society, Inc.

  2. Achilles Tendon Open Surgical Treatment With Platelet-Rich Fibrin Matrix Augmentation: Biomechanical Evaluation.

    Science.gov (United States)

    Alviti, Federica; Gurzì, Michele; Santilli, Valter; Paoloni, Marco; Padua, Roberto; Bernetti, Andrea; Bernardi, Marco; Mangone, Massimiliano

    The relationship between surgical technique and ankle biomechanical properties after surgery for acute rupture of the Achilles tendon (ATR) has not yet been fully investigated. Platelet-rich fibrin (PRF) matrices seem to play a central role in the complex processes of tendon healing. Our aim was to analyze the biomechanical characteristics, stiffness, and mechanical work of the ankle during walking in patients who had undergone surgery after ATR with and without PRF augmentation. We performed a retrospective review of all consecutive patients who had been treated with surgical repair after ATR. Of the 20 male subjects enrolled, 9 (45%) had undergone conventional open repair of the Achilles tendon using the Krackow technique (no-PRF) and 11 (55%) had undergone surgery with PRF augmentation. An additional 8 healthy subjects were included as a control group. A gait analysis evaluation was performed at 6 months after surgery. The percentage of the stance time of the operated leg, double-support time of the healthy leg, and net work of the ankle during the gait cycle showed statistically significant differences between the no-PRF and the healthy group (p < .005). No differences were found between the PRF and healthy groups. Treatment with suture and PRF augmentation could result in significant functional improvements in term of efficiency of motion. Copyright © 2017 American College of Foot and Ankle Surgeons. Published by Elsevier Inc. All rights reserved.

  3. Efficacy of platelet-rich fibrin vs. enamel matrix derivative in the treatment of periodontal intrabony defects: a clinical and cone beam computed tomography study.

    Science.gov (United States)

    Gupta, Swyeta Jain; Jhingran, Rajesh; Gupta, Vivek; Bains, Vivek Kumar; Madan, Rohit; Rizvi, Iram

    2014-07-01

    To evaluate and compare the efficacy of platelet-rich fibrin (PRF) with enamel matrix derivative (EMD; Emdogain) in the treatment of periodontal intrabony defects in patients with chronic periodontitis, six months after surgery. Forty-four (44) intrabony defects in 30 patients (15 males) were randomly allocated into two treatment groups: EMD (n = 22) and PRF (n = 22). Measurement of the defects was done using clinical and cone beam computed tomography at baseline and 6 months. Clinical and radiographic parameters such as probing depth, clinical attachment level, intrabony defect depth and defect angle, were recorded at baseline and 6 months post-operatively. Within group change was evaluated using the Wilcoxon signed rank test. Intergroup comparisons were made using the Mann-Whitney U test. Postsurgical measurements revealed that there was an equal reduction in probing depth and a greater but statistically non-significant attachment gain for the Emdogain group when compared to the platelet-rich fibrin group. The Emdogain group presented with significantly greater percentage defect resolution (43.07% ± 12.21) than did the platelet-rich fibrin group (32.41% ± 14.61). Post-operatively the changes in defect width and defect angle were significant in both groups, but upon intergroup comparison they were found to be statistically non-significantly different. Both Emdogain and platelet-rich fibrin were effective in the regeneration of intrabony defects. Emdogain was significantly superior in terms of percentage defect resolution.

  4. Effect of fibrin glue on the biomechanical properties of human Descemet's membrane.

    Directory of Open Access Journals (Sweden)

    Shyam S Chaurasia

    Full Text Available BACKGROUND: Corneal transplantation has rapidly evolved from full-thickness penetrating keratoplasty (PK to selective tissue corneal transplantation, where only the diseased portions of the patient's corneal tissue are replaced with healthy donor tissue. Descemet's membrane endothelial keratoplasty (DMEK performed in patients with corneal endothelial dysfunction is one such example where only a single layer of endothelial cells with its basement membrane (10-15 µm in thickness, Descemet's membrane (DM is replaced. It is challenging to replace this membrane due to its intrinsic property to roll in an aqueous environment. The main objective of this study was to determine the effects of fibrin glue (FG on the biomechanical properties of DM using atomic force microscopy (AFM and relates these properties to membrane folding propensity. METHODOLOGY/PRINCIPAL FINDINGS: Fibrin glue was sprayed using the EasySpray applicator system, and the biomechanical properties of human DM were determined by AFM. We studied the changes in the "rolling up" tendency of DM by examining the changes in the elasticity and flexural rigidity after the application of FG. Surface topography was assessed using scanning electron microscopy (SEM and AFM imaging. Treatment with FG not only stabilized and stiffened DM but also led to a significant increase in hysteresis of the glue-treated membrane. In addition, flexural or bending rigidity values also increased in FG-treated membranes. CONCLUSIONS/SIGNIFICANCE: Our results suggest that fibrin glue provides rigidity to the DM/endothelial cell complex that may aid in subsequent manipulation by maintaining tissue integrity.

  5. Effect of Fibrin Glue on the Biomechanical Properties of Human Descemet's Membrane

    Science.gov (United States)

    Chaurasia, Shyam S.; Champakalakshmi, Ravi; Li, Ang; Poh, Rebekah; Tan, Xiao Wei; Lakshminarayanan, Rajamani; Lim, Chwee T.; Tan, Donald T.; Mehta, Jodhbir S.

    2012-01-01

    Background Corneal transplantation has rapidly evolved from full-thickness penetrating keratoplasty (PK) to selective tissue corneal transplantation, where only the diseased portions of the patient's corneal tissue are replaced with healthy donor tissue. Descemet's membrane endothelial keratoplasty (DMEK) performed in patients with corneal endothelial dysfunction is one such example where only a single layer of endothelial cells with its basement membrane (10–15 µm in thickness), Descemet's membrane (DM) is replaced. It is challenging to replace this membrane due to its intrinsic property to roll in an aqueous environment. The main objective of this study was to determine the effects of fibrin glue (FG) on the biomechanical properties of DM using atomic force microscopy (AFM) and relates these properties to membrane folding propensity. Methodology/Principal Findings Fibrin glue was sprayed using the EasySpray applicator system, and the biomechanical properties of human DM were determined by AFM. We studied the changes in the “rolling up” tendency of DM by examining the changes in the elasticity and flexural rigidity after the application of FG. Surface topography was assessed using scanning electron microscopy (SEM) and AFM imaging. Treatment with FG not only stabilized and stiffened DM but also led to a significant increase in hysteresis of the glue-treated membrane. In addition, flexural or bending rigidity values also increased in FG-treated membranes. Conclusions/Significance Our results suggest that fibrin glue provides rigidity to the DM/endothelial cell complex that may aid in subsequent manipulation by maintaining tissue integrity. PMID:22662156

  6. The combination use of platelet-rich fibrin and treated dentin matrix for tooth root regeneration by cell homing.

    Science.gov (United States)

    Ji, Baohui; Sheng, Lei; Chen, Gang; Guo, Shujuan; Xie, Li; Yang, Bo; Guo, Weihua; Tian, Weidong

    2015-01-01

    Endogenous regeneration through cell homing provides an alternative approach for tissue regeneration, except cell transplantation, especially considering clinical translation. However, tooth root regeneration through cell homing remains a provocative approach in need of intensive study. Both platelet-rich fibrin (PRF) and treated dentin matrix (TDM) are warehouses of various growth factors, which can promote cell homing. We hypothesized that endogenous stem cells are able to sense biological cues from PRF membrane and TDM, and contribute to the regeneration of tooth root, including soft and hard periodontal tissues. Therefore, the biological effects of canine PRF and TDM on periodontal ligament stem cells (PDLSCs) and bone marrow mesenchymal stem cells (BMSCs) were evaluated respectively in vitro. Beagle dogs were used as orthotopic transplantation model. It was found that PRF significantly recruited and stimulated the proliferation of PDLSCs and BMSCs in vitro. Together, PRF and TDM induced cell differentiation by upregulating the mineralization-related gene expression of bone sialoprotein (BSP) and osteopotin (OPN) after 7 days coculture. In vivo, transplantation of autologous PRF and allogeneic TDM into fresh tooth extraction socket achieved successful root regeneration 3 months postsurgery, characterized by the regeneration of cementum and periodontal ligament (PDL)-like tissues with orientated fibers, indicative of functional restoration. The results suggest that tooth root connected to the alveolar bone by cementum-PDL complex can be regenerated through the implantation of PRF and TDM in a tooth socket microenvironment, probably by homing of BMSCs and PDLSCs. Furthermore, bioactive cues and inductive microenvironment are key factors for endogenous regeneration. This approach provides a tangible pathway toward clinical translation.

  7. Combination of heterologous fibrin sealant and bioengineered human embryonic stem cells to improve regeneration following autogenous sciatic nerve grafting repair.

    Science.gov (United States)

    Mozafari, Roghayeh; Kyrylenko, Sergiy; Castro, Mateus Vidigal; Ferreira, Rui Seabra; Barraviera, Benedito; Oliveira, Alexandre Leite Rodrigues

    2018-01-01

    Peripheral nerve injury is a worldwide clinical problem, and the preferred surgical method for treating it is the end-to-end neurorrhaphy. When it is not possible due to a large nerve gap, autologous nerve grafting is used. However, these surgical techniques result in nerve regeneration at highly variable degrees. It is thus very important to seek complementary techniques to improve motor and sensory recovery. One promising approach could be cell therapy. Transplantation therapy with human embryonic stem cells (hESCs) is appealing because these cells are pluripotent and can differentiate into specialized cell types and have self-renewal ability. Therefore, the main objective of this study was to find conditions under which functional recovery is improved after sciatic nerve neurorrhaphy. We assumed that hESC, either alone or in combination with heterologous fibrin sealant scaffold, could be used to support regeneration in a mouse model of sciatic nerve injury and repair via autografting with end-to-end neurorrhaphy. Five millimeters of the sciatic nerve of C57BL/6 J mice were transected off and rotated 180 degrees to simulate an injury, and then stumps were sutured. Next, we applied heterologous fibrin sealant and/or human embryonic stem cells genetically altered to overexpress fibroblast growth factor 2 (FGF2) at the site of the injury. The study was designed to include six experimental groups comprising neurorrhaphy (N), neurorrhaphy + heterologous fibrin sealant (N + F), neurorrhaphy + heterologous fibrin sealant + doxycycline (N + F + D), neurorrhaphy + heterologous fibrin sealant + wild-type hESC (N + F + W), neurorrhaphy + heterologous fibrin sealant + hESC off (N + F + T), and neurorrhaphy + heterologous fibrin sealant + hESC on via doxycycline (N + F + D + T). We evaluated the recovery rate using Catwalk and von Frey functional recovery tests, as well as immunohistochemistry analysis. The experiments indicated that

  8. A comparative in vitro study of the viability of human keratinocytes grown on irradiated human amnion membrane and fibrin glue scaffolds

    International Nuclear Information System (INIS)

    Dorai, A.A.; Lim, C.K.; Azman, W.S.; Halim, A.S.

    2008-01-01

    Full text: The dried irradiated human amnion membrane has been used as a biological dressing for various clinical conditions. Being another biological membrane its potential as a scaffold to grow human keratinocytes is not known yet. To compare the growth patterns and cell viability of keratinocytes using fibrin glue and air dried amnion membrane as a scaffold. Keratinocytes were obtained from skin samples of six patients undergoing elective surgery. Fibrin glue (Tisseel, Baxter ) was diluted and used to coat the wells. Human dried amnion membrane was obtained and placed into the 24 well plates. Keratinocytes were seeded into the fibrin and amnion scaffold. Cell viability assay (MTT) was performed after 24, 48 and 72 hours. Finally the measurements were done by the Enzyme-Linked Immunosorbent Assay (ELISA) reader at 570 nm. Six patients consented for the study. The cells growing on the amnion scaffold showed a decreasing trend (20.67%, 17.94% and 16.78% respectively for 24, 48 and 72 hours). The cells growing on the fibrin scaffold showed a steady increase in number at 24, 48 and 72 hours (73.03%, 74.12% and 79.66%). The percentage of growth of normal human keratinocytes were significantly greater in the fibrin scaffold group (Mann - Whitney p = 0.002) for 24, 48 and 72 hours. The air dried irradiated human amnion membrane can be used as a scaffold to grow keratinocytes but however the growth pattern does not sustain with time. Fibrin glue supports the growth of human keratinocytes and shows an increasing pattern of growth with time. (Author)

  9. Improved adhesion and differentiation of endothelial cells on surface-attached fibrin structures containing extracellular matrix proteins

    Czech Academy of Sciences Publication Activity Database

    Filová, Elena; Brynda, Eduard; Riedel, Tomáš; Chlupáč, Jaroslav; Vandrovcová, Marta; Švindrych, Zdeněk; Lisá, Věra; Houska, Milan; Pirk, J.; Bačáková, Lucie

    2014-01-01

    Roč. 102, č. 3 (2014), s. 698-712 ISSN 1549-3296 R&D Projects: GA MZd(CZ) NT11270 Institutional support: RVO:67985823 ; RVO:61389013 Keywords : two-dimensional assembly * fibrin * endothelial cells Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery; CD - Macromolecular Chemistry (UMCH-V) Impact factor: 3.369, year: 2014

  10. In-vitro-Studie zum Einfluss von Fibrin in Knorpelkonstrukten auf der Basis von PGA-Vliesstoffen

    DEFF Research Database (Denmark)

    Schmal, H; Mehlhorn, A T; Kurze, C

    2008-01-01

    BACKGROUND: The matrix component in autologous chondrocyte implantation plays an important role. In this study the influence of an additional fibrin component in cartilage constructs based on polyglycolide polymers (PGA) was investigated. METHODS: Human chondrocytes of femoral heads were isolated....... CONCLUSION: Cartilage constructs based on carbohydrate matrices are suitable for matrix-associated chondrocyte implantation. The results of this study suggest a partially inhibitory effect of an additional fibrin component in PGA constructs for chondrogenic differentiation....

  11. Combination of Mineral Trioxide Aggregate and Platelet-rich Fibrin Promotes the Odontoblastic Differentiation and Mineralization of Human Dental Pulp Cells via BMP/Smad Signaling Pathway.

    Science.gov (United States)

    Woo, Su-Mi; Kim, Won-Jae; Lim, Hae-Soon; Choi, Nam-Ki; Kim, Sun-Hun; Kim, Seon-Mi; Jung, Ji-Yeon

    2016-01-01

    Recent reports have shown that the combined use of platelet-rich fibrin (PRF), an autologous fibrin matrix, and mineral trioxide aggregate (MTA) as root filling material is beneficial for the endodontic management of an open apex. However, the potential of the combination of MTA and PRF as an odontogenic inducer in human dental pulp cells (HDPCs) in vitro has not yet been studied. The purpose of this study was to evaluate the effect of the combination of MTA and PRF on odontoblastic maturation in HDPCs. HDPCs extracted from third molars were directly cultured with MTA and PRF extract (PRFe). Odontoblastic differentiation of HDPCs was evaluated by measuring the alkaline phosphatase (ALP) activity, and the expression of odontogenesis-related genes was detected using reverse-transcription polymerase chain reaction or Western blot. Mineralization formation was assessed by alizarin red staining. HDPCs treated with MTA and PRFe significantly up-regulated the expression of dentin sialoprotein and dentin matrix protein-1 and enhanced ALP activity and mineralization compared with those with MTA or PRFe treatment alone. In addition, the combination of MTA and PRFe induced the activation of bone morphogenic proteins (BMP)/Smad, whereas LDN193189, the bone morphogenic protein inhibitor, attenuated dentin sialophosphoprotein and dentin matrix protein-1 expression, ALP activity, and mineralization enhanced by MTA and PRFe treatment. This study shows that the combination of MTA and PRF has a synergistic effect on the stimulation of odontoblastic differentiation of HDPCs via the modulation of the BMP/Smad signaling pathway. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  12. Platelet-Rich Fibrin Lysate Can Ameliorate Dysfunction of Chronically UVA-Irradiated Human Dermal Fibroblasts.

    Science.gov (United States)

    Wirohadidjojo, Yohanes Widodo; Budiyanto, Arief; Soebono, Hardyanto

    2016-09-01

    To determine whether platelet-rich fibrin lysate (PRF-L) could restore the function of chronically ultraviolet-A (UVA)-irradiated human dermal fibroblasts (HDFs), we isolated and sub-cultured HDFs from six different human foreskins. HDFs were divided into two groups: those that received chronic UVA irradiation (total dosages of 10 J cm⁻²) and those that were not irradiated. We compared the proliferation rates, collagen deposition, and migration rates between the groups and between chronically UVA-irradiated HDFs in control and PRF-L-treated media. Our experiment showed that chronic UVA irradiation significantly decreased (p<0.05) the proliferation rates, migration rates, and collagen deposition of HDFs, compared to controls. Compared to control media, chronically UVA-irradiated HDFs in 50% PRF-L had significantly increased proliferation rates, migration rates, and collagen deposition (p<0.05), and the migration rates and collagen deposition of chronically UVA-irradiated HDFs in 50% PRF-L were equal to those of normal fibroblasts. Based on this experiment, we concluded that PRF-L is a good candidate material for treating UVA-induced photoaging of skin, although the best method for its clinical application remains to be determined.

  13. A New Method to Develop Human Dental Pulp Cells and Platelet-rich Fibrin Complex.

    Science.gov (United States)

    He, Xuan; Chen, Wen-Xia; Ban, Guifei; Wei, Wei; Zhou, Jun; Chen, Wen-Jin; Li, Xian-Yu

    2016-11-01

    Platelet-rich fibrin (PRF) has been used as a scaffold material in various tissue regeneration studies. In the previous methods to combine seed cells with PRF, the structure of PRF was damaged, and the manipulation time in vitro was also increased. The objective of this in vitro study was to explore an appropriate method to develop a PRF-human dental pulp cell (hDPC) complex to maintain PRF structure integrity and to find out the most efficient part of PRF. The PRF-hDPC complex was developed at 3 different time points during PRF preparation: (1) the before centrifugation (BC) group, the hDPC suspension was added to the venous blood before blood centrifugation; (2) the immediately after centrifugation (IAC) group, the hDPC suspension was added immediately after blood centrifugation; (3) the after centrifugation (AC) group, the hDPC suspension was added 10 minutes after blood centrifugation; and (4) the control group, PRF without hDPC suspension. The prepared PRF-hDPC complexes were cultured for 7 days. The samples were fixed for histologic, immunohistochemistry, and scanning electron microscopic evaluation. Real-time polymerase chain reaction was performed to evaluate messenger RNA expression of alkaline phosphatase and dentin sialophosphoprotein. Enzyme-linked immunosorbent assay quantification for growth factors was performed within the different parts of the PRF. Histologic, immunohistochemistry, and scanning electron microscopic results revealed that hDPCs were only found in the BC group and exhibited favorable proliferation. Real-time polymerase chain reaction revealed that alkaline phosphatase and dentin sialophosphoprotein expression increased in the cultured PRF-hDPC complex. The lower part of the PRF released the maximum quantity of growth factors. Our new method to develop a PRF-hDPCs complex maintained PRF structure integrity. The hDPCs were distributed in the buffy coat, which might be the most efficient part of PRF. Copyright © 2016 American

  14. Comparing different preparation methods to study human fibrin fibers and platelets using TEM.

    Science.gov (United States)

    Buys, Antoinette V; Pretorius, Etheresia

    2012-06-01

    For the study of cellular ultrastructure, the sample needs to be stabilized by fixation, with the ultimate aim to preserve the native tissue organization and to protect the tissue against later stages of preparation. Chemical and freezing fixation are most used, and chemical fixation employs agents that permeate tissues and cells by diffusion and covalently bind with their major biochemical constituents to fix them. Most widely used chemical fixatives are aldehydes, e.g., formaldehyde and glutaraldehyde, which are noncoagulating, crosslinking agents. Cryofixation methods for ultrastructural studies are also popular, and high-pressure freezing immobilizes all cell constituents and arrests biological activity by removing the thermal energy from the system. In the current research, we used platelet-rich plasma (PRP) to study expansive fibrin fibers and platelet ultrastructure to compare the two fixation techniques. We also used thrombin and calcium chloride as a clotting agent to determine the technique most suitable for the formation of extensive fibrin networks. Chemically fixated fibrin fibers were more compact and condensed and also showed a banding pattern on longitudinal sections. High-pressure frozen samples were more dispersed while platelets fixated showed better preserved cellular membranes and organelle structure. PRP coagulated by addition of CaCl(2) showed blood platelets that are noticeably more activated compared with PRP; however, with thrombin, a sharp ultrastructure was seen. We conclude that PRP mixed with thrombin, and freeze substituted, is the most suitable method for the study of extensive fibrin fibers as well as platelets. Copyright © 2011 Wiley Periodicals, Inc.

  15. Novel use of platelet-rich fibrin matrix and MTA as an apical barrier in the management of a failed revascularization case.

    Science.gov (United States)

    Yadav, Pankaj; Pruthi, Preeti Jain; Naval, Ruchika Roongta; Talwar, Sangeeta; Verma, Mahesh

    2015-08-01

    We report management of a failed revascularization/revitalization case, which could be due to inadequate removal of biofilm and bacteria in dentinal tubules. The use of an apical matrix barrier in form of a platelet-rich fibrin (PRF) membrane for stabilization of MTA in root end apexification procedure is described. The canal was cleansed of old MTA present in the cervical third using H files, irrigated using saline and finally irrigated with 2.5% NaOCl and saline. To obtain canal disinfection, calcium hydroxide paste was temporized in the canal. In subsequent appointments, PRF was placed at the root tip followed by 5-mm apical plug with mineral trioxide aggregate. One week later, the root canal was obturated with thermoplasticized gutta-percha. A 6-month and a 2-year follow ups showed reduction of periapical radiolucency and adequately functional tooth. One-visit apexification techniques provide an alternative treatment for failed revascularization cases. Follow up confirmed complete healing periradicularly. Apexification in one step using an apical barrier of PRF and a plug of MTA can be considered a predictable treatment and may be an alternative to long-term revascularization failures. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  16. Effects of Plasma Rich in Growth Factors and Platelet-Rich Fibrin on Proliferation and Viability of Human Gingival Fibroblasts

    Science.gov (United States)

    Vahabi, Surena; Vaziri, Shahram; Torshabi, Maryam

    2015-01-01

    Objectives: Platelet preparations are commonly used to enhance bone and soft tissue regeneration. Considering the existing controversies on the efficacy of platelet products for tissue regeneration, more in vitro studies are required. The aim of the present study was to compare the in vitro effects of plasma rich in growth factors (PRGF) and platelet-rich fibrin (PRF) on proliferation and viability of human gingival fibroblasts (HGFs). Materials and Methods: Anitua’s PRGF and Choukran’s PRF were prepared according to the standard protocols. After culture periods of 24, 48 and 72 hours, proliferation of HGFs was evaluated by the methyl thiazol tetrazolium assay. Statistical analysis was performed using one-way ANOVA followed by Tukey-Kramer’s multiple comparisons and P-valuesPRGF treatment induced statistically significant (PPRGF than in PRF group (PPRGF had a strong stimulatory effect on HGF cell viability and proliferation compared to PRF. PMID:26877740

  17. A review of fibrin and fibrin composites for bone tissue engineering.

    Science.gov (United States)

    Noori, Alireza; Ashrafi, Seyed Jamal; Vaez-Ghaemi, Roza; Hatamian-Zaremi, Ashraf; Webster, Thomas J

    2017-01-01

    Tissue engineering has emerged as a new treatment approach for bone repair and regeneration seeking to address limitations associated with current therapies, such as autologous bone grafting. While many bone tissue engineering approaches have traditionally focused on synthetic materials (such as polymers or hydrogels), there has been a lot of excitement surrounding the use of natural materials due to their biologically inspired properties. Fibrin is a natural scaffold formed following tissue injury that initiates hemostasis and provides the initial matrix useful for cell adhesion, migration, proliferation, and differentiation. Fibrin has captured the interest of bone tissue engineers due to its excellent biocompatibility, controllable biodegradability, and ability to deliver cells and biomolecules. Fibrin is particularly appealing because its precursors, fibrinogen, and thrombin, which can be derived from the patient's own blood, enable the fabrication of completely autologous scaffolds. In this article, we highlight the unique properties of fibrin as a scaffolding material to treat bone defects. Moreover, we emphasize its role in bone tissue engineering nanocomposites where approaches further emulate the natural nanostructured features of bone when using fibrin and other nanomaterials. We also review the preparation methods of fibrin glue and then discuss a wide range of fibrin applications in bone tissue engineering. These include the delivery of cells and/or biomolecules to a defect site, distributing cells, and/or growth factors throughout other pre-formed scaffolds and enhancing the physical as well as biological properties of other biomaterials. Thoughts on the future direction of fibrin research for bone tissue engineering are also presented. In the future, the development of fibrin precursors as recombinant proteins will solve problems associated with using multiple or single-donor fibrin glue, and the combination of nanomaterials that allow for the

  18. A review of fibrin and fibrin composites for bone tissue engineering

    Directory of Open Access Journals (Sweden)

    Noori A

    2017-07-01

    Full Text Available Alireza Noori,1 Seyed Jamal Ashrafi,2 Roza Vaez-Ghaemi,3 Ashraf Hatamian-Zaremi,4 Thomas J Webster5 1Department of Tissue Engineering and Applied Cell Sciences, Faculty of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, 2School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran; 3Department of Chemical and Biological Engineering, Faculty of Biomedical Engineering, The University of British Columbia, Vancouver, BC, Canada; 4Faculty of New Sciences and Technologies, University of Tehran, Tehran, Iran; 5Department of Chemical Engineering, Northeastern University, Boston, MA, USA Abstract: Tissue engineering has emerged as a new treatment approach for bone repair and regeneration seeking to address limitations associated with current therapies, such as autologous bone grafting. While many bone tissue engineering approaches have traditionally focused on synthetic materials (such as polymers or hydrogels, there has been a lot of excitement surrounding the use of natural materials due to their biologically inspired properties. Fibrin is a natural scaffold formed following tissue injury that initiates hemostasis and provides the initial matrix useful for cell adhesion, migration, proliferation, and differentiation. Fibrin has captured the interest of bone tissue engineers due to its excellent biocompatibility, controllable biodegradability, and ability to deliver cells and biomolecules. Fibrin is particularly appealing because its precursors, fibrinogen, and thrombin, which can be derived from the patient’s own blood, enable the fabrication of completely autologous scaffolds. In this article, we highlight the unique properties of fibrin as a scaffolding material to treat bone defects. Moreover, we emphasize its role in bone tissue engineering nanocomposites where approaches further emulate the natural nanostructured features of bone when using fibrin and other nanomaterials. We also review the

  19. Experimental Evaluation of the Effectiveness of Demineralized Bone Matrix and Collagenated Heterologous Bone Grafts Used Alone or in Combination with Platelet-Rich Fibrin on Bone Healing in Sinus Floor Augmentation.

    Science.gov (United States)

    Peker, Elif; Karaca, Inci Rana; Yildirim, Benay

    2016-01-01

    The aim of this study was an experimental evaluation of the effectiveness of demineralized bone matrix (DBM) and collagenated heterologous bone graft (CHBG) used alone or in combination with platelet-rich fibrin on bone healing in sinus floor augmentation procedures. In this study, 36 New Zealand rabbits were used. The bilateral sinus elevation was performed, and 72 defects were obtained. The rabbit maxillary sinuses were divided into four groups according to the augmentation biomaterials obtained: demineralized bone matrix (Grafton DBM Putty, Osteotech; DBM group), DBM combined with platelet-rich fibrin (PRF; DBM + PRF group), collagenated heterologous bone graft (CHBG; Apatos Mix, OsteoBiol, Tecnoss; CHBG group), CHBG combined with PRF (CHBG + PRF group). All groups were sacrificed at 2, 4, and 8 weeks after surgery for histologic, histomorphometric, and immunohistochemical analyses. The inflammatory reaction was moderate to intense at the second week in all groups and declined from 2 to 8 weeks. New bone formation was started at the second week and increased from 2 to 8 weeks in all groups. There was no significant difference in bone formation between the experimental groups that used PRF mixed graft material and control groups that used only graft material. The percentage of new bone formation showed a significant difference in DBM groups and DBM + PRF groups compared with other groups. There were osteoclasts around all the bone graft materials used, but the percentage of residual graft particles was significantly higher in CHBG groups and CHBG + PRF groups at the eighth week. There is no beneficial effect of the application of PRF in combination with demineralized bone matrix or collagenated heterologous bone graft on bone formation in sinus floor augmentation. The results of this study showed that both collagenated heterologous bone graft and demineralized bone matrix have osteoconductive properties, but demineralized bone matrix showed more bone formation

  20. Availability of the B beta(15-21) epitope on cross-linked human fibrin and its plasmic degradation products

    Science.gov (United States)

    Chen, F.; Haber, E.; Matsueda, G. R.

    1992-01-01

    The binding of radiolabeled monoclonal antifibrin antibody 59D8 (specific for fibrin but not fibrinogen) to a series of degraded fibrin clots showed that the availability of the B beta(15-21) epitope (against which 59D8 had been raised) was inversely proportional to the extent of clot lysis. Examination of digest supernatants revealed that the B beta(15-21) epitope was released from clots as a high molecular weight degradation product in the presence of calcium ions but that the generation of low molecular weight peptides occurred in the absence of calcium ions. To address the question of epitope accessibility, we compared levels of fibrin clot binding among four radioactively labeled antibodies: antifibrin monoclonal antibody 59D8, two antifibrinogen monoclonal antibodies that cross-reacted with fibrin, and an affinity-purified polyclonal antifibrinogen antibody. We expected that the antifibrinogen antibodies would show enhanced binding to clots in comparison with the antifibrin antibody. However, the epitope accessibility experiments showed that all four antibody preparations bound fibrin clots at comparable levels. Taken together, these studies demonstrated that one fibrin-specific epitope, B beta(15-21), remains available on clots as they undergo degradation by plasmin and, importantly, that the epitope is not solubilized at a rate faster than the rate at which the clot is itself solubilized. The availability of the B beta(15-21) epitope during the course of plasminolysis assures the potential utility of antifibrin antibodies such as 59D8 for detecting thrombi and targeting plasminogen activators.

  1. Biomimetic chimeric peptide-tethered hydrogels for human mesenchymal stem cell delivery.

    Science.gov (United States)

    Shim, Gayong; Kim, Gunwoo; Choi, Junhyeok; Yi, TacGhee; Cho, Yun Kyoung; Song, Sun Uk; Byun, Youngro; Oh, Yu-Kyoung

    2015-12-01

    Here, we report a chimeric peptide-tethered fibrin hydrogel scaffold for delivery of human mesenchymal stem cells (hMSC). Osteopontin-derived peptide (OP) was used as an hMSC-tethering moiety. OP showed hMSC adhesion properties and enhanced hMSC proliferation. A natural fibrin-binding protein-derived peptide (FBP) was tested for its ability to tether hMSC to the fibrin gel matrix. FBP loading on fibrin gels was 8.2-fold higher than that of a scrambled peptide (scFBP). FBP-loaded fibrin gels were retained at injection sites longer than scFBP-loaded fibrin gels, showing a 15.9-fold higher photon intensity of fluorescent FBP-grafted fibrin gels than fluorescent scFBP-loaded fibrin gels 48 h after injection. On the basis of the fibrin gel-binding properties of FBP and the hMSC-binding and proliferation-supporting properties of OP, we constructed chimeric peptides containing FBP and OP linked with a spacer (FBPsOP). Four days after transplantation, the survival of hMSC in FBPsOP-grafted fibrin gels was 3.9-fold higher than hMSC in fibrin gels alone. Our results suggest the potential of FBPsOP-grafted fibrin gels as a bioactive delivery system for enhanced survival of stem cells. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Fibrin(ogen) is internalized and degraded by activated human monocytoid cells via Mac-1 (CD11b/CD18): a nonplasmin fibrinolytic pathway.

    Science.gov (United States)

    Simon, D I; Ezratty, A M; Francis, S A; Rennke, H; Loscalzo, J

    1993-10-15

    Fibrin(ogen) (FGN) is important for hemostasis and wound healing and is cleared from sites of injury primarily by the plasminogen activator system. However, there is emerging evidence in plasminogen activator-deficient transgenic mice that nonplasmin pathways may be important in fibrin(ogen)olysis, as well. Given the proximity of FGN and monocytes within the occlusive thrombus at sites of vascular injury, we considered the possibility that monocytes may play an ancillary role in the degradation and clearance of fibrin. We found that monocytes possess an alternative fibrinolytic pathway that uses the integrin Mac-1, which directly binds and internalizes FGN, resulting in its lysosomal degradation. At 4 degrees C, FGN binds to U937 monocytoid cells in a specific and saturable manner with a kd of 1.8 mumol/L. Binding requires adenosine diphosphate stimulation and is calcium-dependent. At 37 degrees C, FGN and fibrin monomer (FM) are internalized and degraded at rates of 0.37 +/- 0.13 and 0.55 +/- 0.03 microgram/10(6) cells/h by U937 cells, 1.38 +/- 0.02 and 1.20 +/- 0.30 microgram/10(6) cells/h by THP-1 cells, and 2.10 +/- 0.20 and 2.52 +/- 0.18 micrograms/10(6) cells/h by human peripheral blood mononuclear cells, respectively. The serine protease inhibitors, PPACK and aprotinin, and the specific elastase inhibitor, AAPVCK, do not significantly inhibit degradation. However, degradation is inhibited by chloroquine, suggesting that a lysosomal pathway is involved. Factor X, a competitive ligand with FGN for the Mac-1 receptor, also blocks degradation, as does a monoclonal antibody to the alpha-subunit of Mac-1. Autoradiography of radioiodinated, internalized FGN shows that FGN proteolysis by the pathway produces a unique degradation pattern distinct from that observed with plasmin. In a fibrin clot lysis assay, Mac-1-mediated fibrinolysis contributed significantly to total fibrinolysis. In summary, FGN is internalized and degraded by activated human monocytoid cells via

  3. FIBRIN-TYPE FIBRINOID IN HUMAN PLACENTA: A STEREOLOGICAL ANALYSIS OF ITS ASSOCIATION WITH INTERVILLOUS VOLUME AND VILLOUS SURFACE AREA

    Directory of Open Access Journals (Sweden)

    Terry M Mayhew

    2011-05-01

    Full Text Available Stereological methods were used to examine fibrin-type fibrinoid deposition in the intervillous spaces of human placentas collected during gestation (12-41 weeks and from term pregnancies at low (400 m and high (3.6 km altitude. The main aim was to test predictions about the relationships between fibrinoid deposits and either the volume of intervillous space or the surface area of (intermediate + terminal villi. Fields of view on Masson trichrome-stained paraffin sections were selected as part of a systematic sampling design which randomised section location and orientation. Relative and absolute volumes were estimated by test point counting and surfaces by intersection counting. Apparent differences were tested by analyses of variance and relationships by correlation and regression analysis. Fibrinoid volume increased during gestation and correlated positively with intervillous volume and villous surface area. However, relative to intervillous volume, the main increase in fibrinoid occurred towards term (36-41 weeks. At high altitude, placentas contained more intervillous space but less fibrinoid. At both altitudes, there were significant correlations between fibrinoid volume and villous surface area. In all cases, changes in fibrinoid volume were commensurate with changes in villous surface area. Whilst findings lend support to the notion that fibrinoid deposition during normal gestation is influenced by the quality of vascular perfusion, they also emphasise that the extent of the villous surface is a more generally important factor. The villous surface may influence the steady state between coagulation and fibrinolysis since some pro-coagulatory events operate at the trophoblastic epithelium. They occur notably at sites of trophoblast de-epithelialisation and these arise following trauma or during the extrusion phase of normal epithelial turnover.

  4. Effects of Plasma Rich in Growth Factors and Platelet-Rich Fibrin on Proliferation and Viability of Human Gingival Fibroblasts

    Directory of Open Access Journals (Sweden)

    Surena Vahabi

    2016-01-01

    Full Text Available Objectives: Platelet preparations are commonly used to enhance bone and soft tissue regeneration. Considering the existing controversies on the efficacy of platelet products for tissue regeneration, more in vitro studies are required. The aim of the present study was to compare the in vitro effects of plasma rich in growth factors (PRGF and platelet-rich fibrin (PRF on proliferation and viability of human gingival fibroblasts (HGFs.Materials and Methods: Anitua's PRGF and Choukran's PRF were prepared according to the standard protocols. After culture periods of 24, 48 and 72 hours, proliferation of HGFs was evaluated by the methyl thiazol tetrazolium assay. Statistical analysis was performed using one-way ANOVA followed by Tukey-Kramer’s multiple comparisons and P-values<0.05 were considered statistically significant.Results: PRGF treatment induced statistically significant (P<0.001 proliferation of HGF cells compared to the negative control (100% viability at 24, 48 and 72 hours in values of 123%±2.25%, 102%±2.8% and 101%±3.92%, respectively. The PRF membrane treatment of HGF cells had a statistically significant effect on cell proliferation (21%±1.73%, P<0.001 at 24 hours compared to the negative control. However, at 48 and 72 hours after treatment, PRF had a negative effect on HGF cell proliferation and caused 38% and 60% decrease in viability and proliferation compared to the negative control, respectively. The HGF cell proliferation was significantly higher in PRGF than in PRF group (P< 0.001.Conclusion: This study demonstrated that PRGF had a strong stimulatory effect on HGF cell viability and proliferation compared to PRF.

  5. [Research of repairing rabbit knee joint cartilage defect by compound material of fibrin glue and decalcified bone matrix (DBM) and chondrocytes].

    Science.gov (United States)

    He, Jie; Yang, Xiang; Yue, Peng-ju; Wang, Guan-yu; Guo, Ting; Zhao, Jian-ning

    2009-07-01

    To investigate the feasibility and effectivity of using compound material of fibrin glue and DBM as scaffolds for cartilage tissue engineering. Chondrocytes isolated from articular cartilage were seeded into prepared scaffolds, after incubation for 4 weeks in vitro. Chondrocytes and fibrin glue and DBM constructs were implanted in the joint cave of rabbit. The specimens were excised at the 4th, 8th, 12th week, examined grossly analyzed by haematoxylin cosine, toluidine blues staining and type II collagen immunohistochemistry reaction. Wakitani score was counted to evaluate the repairing effect. Grossly analysis showed some ivory tissue filled the caves after 4 weeks and the caves were full filled with smooth surface after 12 weeks. The microscope showed a good deal of chondrocytes appeared after 8 weeks and more type II collagen than 4 weeks. Twelve weeks later, cartilage lacuna could be observed. The cells arrangement and the amount of type II collagen both showed the same as the natural one. Complicated material of fibrin glue and DBM as scaffolds can be used as scaffolds for cartilage tissue engineering.

  6. FIBRIN GLUE DAN APLIKASINYA

    Directory of Open Access Journals (Sweden)

    Agi Harliani S

    2015-08-01

    Full Text Available Fibrin Tissue Adhesive (FTA, Fibrin Sealant (FS or Fibrin Glue (FG are names given to a group of product that lead to the formation of fibrin clot at the site of application. Fibrin Glue represents a new revolution for local haemostatic, which produced by based on the understanding about blood coagulation process. The mechanism of FG mimics the last stage of blood coagulation process. Haemophilia, is a congenital inherited bleeding disorder, characterized by repeated bleeding episodes. The basic pathology is deficiency of factor VIII (hemophilia A or factor IX (hemophilia B. At bleeding episodes, hemophilia patients need replacement therapy. Hemophilia patients need transfusion of cryoprecipitate, Fresh Frozen Plasma (FFP or factor concentrate as replacement therapy. Oral surgery, dental extraction, circumcision, and orthopedic operations are the most important indications for fibrin glue in hemophilia care. As haemostatic local, FG minimizes bleeding, reducing the need of transfusion or factor concentrate, reducing the complication of transfusion, hospitalization and cost.

  7. Optical properties of an anterior lamellar human cornea model based on fibrin-agarose

    Science.gov (United States)

    Ionescu, Ana M.; Cardona, Juan de la Cruz; Ghinea, Razvan; Garzón, Ingrid; González-Andrades, Miguel; Alaminos, Miguel; Pérez, Maria del Mar

    2017-08-01

    The optical evaluation carried out using the Inverse Adding-Doubling (IAD) method to determine the scattering and the absorption coefficients of the bioengineered human corneal stromas showed that this type of artificial biomaterials shared many similarities with native control cornea after four weeks of development in culture. Their absorption and reduced scattering coefficients values were higher than the ones of the control cornea, but their spectral behaviors of both coefficients were similar. Time of development in culture was an influencing factor on the results.

  8. Human Homeostatic Control Matrix in Norm

    Directory of Open Access Journals (Sweden)

    Alexander G. Kruglov

    2016-09-01

    Full Text Available We undertook our research to study and systemize the relationship between hemodynamics and biochemical parameters of arterial and venous blood in healthy people. Hemodynamic and biochemical characteristics were obtained through a probe by using catheterization in various vascular areas (aorta, brain, heart, lungs, and liver. Correlation and factor analyses were conducted to study the relationship between the obtained characteristics of the regional and systemic blood flow. Due to the nature of the correlation analysis, the significant (p<0.05 relation signs (+, 0, - without regard to their power were considered. The obtained results suggested that there are sets of both intra-organ and system regulatory relationships between metabolic and hemodynamic characteristics. The complex of relationships among the studied parameters makes it possible to maintain the homeostatic equilibrium in the body. The psychophysiological control system includes the subsystems we described: 1 the cardiac-hepatic-pulmonary complex having properties of the metabolic and hemodynamic information field providing biological stability of the homeostasis; any significant imbalance of its elements triggers afferent information flows actualizing an afferent synthesis; 2 the mind forming gradient patterns of targeted behavior to eliminate metabolic imbalance, to achieve goals both as coded biological parameters and as the highest forms of behavior, to reach the ultimate goal: parametric, homeostatic equilibrium in the “biosphere” of the human body. By using the results of our research and the complex of dynamic relationships in human homeostasis, we built a homeostatic control matrix (HCM.

  9. The initiation of embryonic-like collagen fibrillogenesis by adult human tendon fibroblasts when cultured under tension

    DEFF Research Database (Denmark)

    Bayer, Monika L; Yeung, Chin-Yan C; Kadler, Karl E

    2010-01-01

    to initiate collagen fibrillogenesis when cultured in fixed-length fibrin gels. Fibroblasts were dissected from semitendinosus and gracilis tendons from healthy humans and cultured in 3D linear fibrin gels. The fibroblasts synthesized an extracellular matrix of parallel collagen fibrils that were aligned...

  10. 21 CFR 864.7320 - Fibrinogen/fibrin degradation products assay.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Fibrinogen/fibrin degradation products assay. 864.7320 Section 864.7320 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN....7320 Fibrinogen/fibrin degradation products assay. (a) Identification. A fibrinogen/fibrin degradation...

  11. 21 CFR 864.7300 - Fibrin monomer paracoagulation test.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Fibrin monomer paracoagulation test. 864.7300 Section 864.7300 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7300 Fibrin...

  12. Characterization of Leukocyte-platelet Rich Fibrin, A Novel Biomaterial

    OpenAIRE

    Madurantakam, Parthasarathy; Yoganarasimha, Suyog; Hasan, Fadi K.

    2015-01-01

    Autologous platelet concentrates represent promising innovative tools in the field of regenerative medicine and have been extensively used in oral surgery. Unlike platelet rich plasma (PRP) that is a gel or a suspension, Leukocyte-Platelet Rich Fibrin (L-PRF) is a solid 3D fibrin membrane generated chair-side from whole blood containing no anti-coagulant. The membrane has a dense three dimensional fibrin matrix with enriched platelets and abundant growth factors. L-PRF is a popular adjunct in...

  13. Random matrix analysis of human EEG data

    Czech Academy of Sciences Publication Activity Database

    Šeba, Petr

    2003-01-01

    Roč. 91, - (2003), s. 198104-1 - 198104-4 ISSN 0031-9007 R&D Projects: GA ČR GA202/02/0088 Institutional research plan: CEZ:AV0Z1010914 Keywords : random matrix theory * EEG signal Subject RIV: BE - Theoretical Physics Impact factor: 7.035, year: 2003

  14. Fibrinogen and fibrin.

    Science.gov (United States)

    Weisel, John W

    2005-01-01

    Fibrinogen is a large, complex, fibrous glycoprotein with three pairs of polypeptide chains linked together by 29 disulfide bonds. It is 45 nm in length, with globular domains at each end and in the middle connected by alpha-helical coiled-coil rods. Both strongly and weakly bound calcium ions are important for maintenance of fibrinogen's structure and functions. The fibrinopeptides, which are in the central region, are cleaved by thrombin to convert soluble fibrinogen to insoluble fibrin polymer, via intermolecular interactions of the "knobs" exposed by fibrinopeptide removal with "holes" always exposed at the ends of the molecules. Fibrin monomers polymerize via these specific and tightly controlled binding interactions to make half-staggered oligomers that lengthen into protofibrils. The protofibrils aggregate laterally to make fibers, which then branch to yield a three-dimensional network-the fibrin clot-essential for hemostasis. X-ray crystallographic structures of portions of fibrinogen have provided some details on how these interactions occur. Finally, the transglutaminase, Factor XIIIa, covalently binds specific glutamine residues in one fibrin molecule to lysine residues in another via isopeptide bonds, stabilizing the clot against mechanical, chemical, and proteolytic insults. The gene regulation of fibrinogen synthesis and its assembly into multichain complexes proceed via a series of well-defined steps. Alternate splicing of two of the chains yields common variant molecular isoforms. The mechanical properties of clots, which can be quite variable, are essential to fibrin's functions in hemostasis and wound healing. The fibrinolytic system, with the zymogen plasminogen binding to fibrin together with tissue-type plasminogen activator to promote activation to the active enzyme plasmin, results in digestion of fibrin at specific lysine residues. Fibrin(ogen) also specifically binds a variety of other proteins, including fibronectin, albumin

  15. Conformational changes in fragments D and double-D from human fibrin(ogen) upon binding the peptide ligand Gly-His-Arg-Pro-amide.

    Science.gov (United States)

    Everse, S J; Spraggon, G; Veerapandian, L; Doolittle, R F

    1999-03-09

    The structure of fragment double-D from human fibrin has been solved in the presence and absence of the peptide ligands that simulate the two knobs exposed by the removal of fibrinopeptides A and B, respectively. All told, six crystal structures have been determined, three of which are reported here for the first time: namely, fragments D and double-D with the peptide GHRPam alone and double-D in the absence of any peptide ligand. Comparison of the structures has revealed a series of conformational changes that are brought about by the various knob-hole interactions. Of greatest interest is a moveable "flap" of two negatively charged amino acids (Glubeta397 and Aspbeta398) whose side chains are pinned back to the coiled coil with a calcium atom bridge until GHRPam occupies the beta-chain pocket. Additionally, in the absence of the peptide ligand GPRPam, GHRPam binds to the gamma-chain pocket, a new calcium-binding site being formed concomitantly.

  16. Ginsenoside Rg1 and platelet-rich fibrin enhance human breast adipose-derived stem cells function for soft tissue regeneration

    Science.gov (United States)

    Li, Hong-Mian; Peng, Qi-Liu; Huang, Min-Hong; Li, De-Quan; Liang, Yi-Dan; Chi, Gang-Yi; Li, De-Hui; Yu, Bing-Chao; Huang, Ji-Rong

    2016-01-01

    Adipose-derived stem cells (ASCs) can be used to repair soft tissue defects, wounds, burns, and scars and to regenerate various damaged tissues. The cell differentiation capacity of ASCs is crucial for engineered adipose tissue regeneration in reconstructive and plastic surgery. We previously reported that ginsenoside Rg1 (G-Rg1 or Rg1) promotes proliferation and differentiation of ASCs in vitro and in vivio. Here we show that both G-Rg1 and platelet-rich fibrin (PRF) improve the proliferation, differentiation, and soft tissue regeneration capacity of human breast adipose-derived stem cells (HBASCs) on collagen type I sponge scaffolds in vitro and in vivo. Three months after transplantation, tissue wet weight, adipocyte number, intracellular lipid, microvessel density, and gene and protein expression of VEGF, HIF-1α, and PPARγ were higher in both G-Rg1- and PRF-treated HBASCs than in control grafts. More extensive new adipose tissue formation was evident after treatment with G-Rg1 or PRF. In summary, G-Rg1 and/or PRF co-administration improves the function of HBASCs for soft tissue regeneration engineering. PMID:27191987

  17. Fibrin glue in ophthalmology

    Directory of Open Access Journals (Sweden)

    Panda Anita

    2009-01-01

    Full Text Available Suturing is a time consuming task in ophthalmology and suture induced irritation and redness are frequent problems. Postoperative wound infection and corneal graft rejection are examples of possible suture related complications. To prevent these complications, ophthalmic surgeons are switching to sutureless surgery. A number of recent developments have established tissue adhesives like cyanoacrylate glue and fibrin glue as attractive alternatives to sutures. A possible and promising new application for tissue adhesives is to provide a platform for tissue engineering. Currently, tissue glue is being used for conjunctival closure following pterygium and strabismus surgery, forniceal reconstruction surgery, amniotic membrane transplantation, lamellar corneal grafting, closure of corneal perforations and descematoceles, management of conjunctival wound leaks after trabeculectomy, lid surgery, adnexal surgery and as a hemostat to minimise bleeding. The purpose of this review is to discuss the currently available information on fibrin glue.

  18. Fibrin glue mixed with platelet-rich fibrin as a scaffold seeded with dental bud cells for tooth regeneration.

    Science.gov (United States)

    Yang, Kai-Chiang; Wang, Chun-Hao; Chang, Hao-Hueng; Chan, Wing P; Chi, Chau-Hwa; Kuo, Tzong-Fu

    2012-11-01

    Odontogenesis is a complex process with a series of epithelial-mesenchymal interactions and odontogenic molecular cascades. In tissue engineering of teeth from stem cells, platelet-rich fibrin (PRF), which is rich in growth factors and cytokines, may improve regeneration. Accordingly, PRF was added into fibrin glue to enrich the microenvironment with growth factors. Unerupted second molar tooth buds were harvested from miniature swine and cultured in vitro for 3 weeks to obtain dental bud cells (DBCs). Whole blood was collected for the preparation of PRF and fibrin glue before surgery. DBCs were suspended in fibrin glue and then enclosed with PRF, and the DBC-fibrin glue-PRF composite was autografted back into the original alveolar sockets. Radiographic and histological examinations were used to identify the regenerated tooth structure 36 weeks after implantation. Immunohistochemical staining was used to detect proteins specific to tooth regeneration. One pig developed a complete tooth with crown, root, pulp, enamel, dentin, odontoblast, cementum, blood vessels, and periodontal ligaments in indiscriminate shape. Another animal had an unerupted tooth that expressed cytokeratin 14, dentin matrix protein-1, vascular endothelial growth factor, and osteopontin. This study demonstrated, using autogenic cell transplantation in a porcine model, that DBCs seeded into fibrin glue-PRF could regenerate a complete tooth. Copyright © 2011 John Wiley & Sons, Ltd.

  19. In vivo extracellular matrix protein expression by human periodontal ...

    African Journals Online (AJOL)

    It is well known that the orthodontic force applied to teeth generates a series of events that remodel the periodontal ligament (PDL). Extracellular matrix proteins (ECM) are described as molecular regulators of these events. However, the exact contribution of these proteins in human PDL modeling by orthodontic force ...

  20. Assessment of cartilage repair after chondrocyte transplantation with a fibrin-hyaluronan matrix – Correlation of morphological MRI, biochemical T2 mapping and clinical outcome

    International Nuclear Information System (INIS)

    Eshed, Iris; Trattnig, Siegfried; Sharon, Michal; Arbel, Ron; Nierenberg, Gabriel; Konen, Eli; Yayon, Avner

    2012-01-01

    Objective: To evaluate change over time of clinical scores, morphological MRI of cartilage appearance and quantitative T2 values after implantation with BioCart™II, a second generation matrix-assisted implantation system. Methods: Thirty-one patients were recruited 6–49 months post surgery for cartilage defect in the femoral condyle. Subjects underwent MRI (morphological and T2-mapping sequences) and completed the International Knee Documentation Committee (IKDC) questionnaire. MRI scans were scored using the MR Observation of Cartilage Repair Tissue (MOCART) system and cartilage T2-mapping values were registered. Analysis included correlation of IKDC scores, MOCART and T2 evaluation with each other, with implant age and with previous surgical intervention history. Results: IKDC score significantly correlated with MOCART score (r = −0.39, p = 0.031), inversely correlated with previous interventions (r = −0.39, p = 0.034) and was significantly higher in patients with longer follow-up time (p = 0.0028). MOCART score was slight, but not significantly higher in patients with longer term implants (p = 0.199). T2 values were significantly lower in patients with longer duration implants (p < 0.001). This trend was repeated in patients with previous interventions, although to a lesser extent. Conclusions: Significant improvement with time from BioCart™II implantation can be expected by IKDC scoring and MRI T2-mapping values. Patients with previous knee operations can also benefit from this procedure.

  1. Composition of fibrin glues significantly influences axial vascularization and degradation in isolation chamber model.

    Science.gov (United States)

    Arkudas, Andreas; Pryymachuk, Galyna; Hoereth, Tobias; Beier, Justus P; Polykandriotis, Elias; Bleiziffer, Oliver; Gulle, Heinz; Horch, Raymund E; Kneser, Ulrich

    2012-07-01

    In this study, different fibrin sealants with varying concentrations of the fibrin components were evaluated in terms of matrix degradation and vascularization in the arteriovenous loop (AVL) model of the rat. An AVL was placed in a Teflon isolation chamber filled with 500 μl fibrin gel. The matrix was composed of commercially available fibrin gels, namely Beriplast (Behring GmbH, Marburg, Germany) (group A), Evicel (Omrix Biopharmaceuticals S.A., Somerville, New Jersey, USA) (group B), Tisseel VH S/D (Baxter, Vienna, Austria) with a thrombin concentration of 4 IU/ml and a fibrinogen concentration of 80 mg/ml [Tisseel S F80 (Baxter), group C] and with an fibrinogen concentration of 20 mg/ml [Tisseel S F20 (Baxter), group D]. After 2 and 4 weeks, five constructs per group and time point were investigated using micro-computed tomography, and histological and morphometrical analysis techniques. The aprotinin, factor XIII and thrombin concentration did not affect the degree of clot degradation. An inverse relationship was found between fibrin matrix degradation and sprouting of blood vessels. By reducing the fibrinogen concentration in group D, a significantly decreased construct weight and an increased generation of vascularized connective tissue were detected. There was an inverse relationship between matrix degradation and vascularization detectable. Fibrinogen as the major matrix component showed a significant impact on the matrix properties. Alteration of fibrin gel properties might optimize formation of blood vessels.

  2. Platelet rich fibrin combined with decalcified freeze-dried bone allograft for the treatment of human intrabony periodontal defects: a randomized split mouth clinical trail.

    Science.gov (United States)

    Agarwal, Ashish; Gupta, Narinder Dev; Jain, Avikal

    2016-01-01

    Polypeptide growth factors of platelet rich fibrin (PRF) have the potential to regenerate periodontal tissues. Osteoinductive property of demineralized freeze-dried bone allograft (DFDBA) has been successfully utilized in periodontal regeneration. The aim of the present randomized, split mouth, clinical trial was to determine the additive effects of PRF with a DFDBA in the treatment of human intrabony periodontal defects. Sixty interproximal infrabony defects in 30 healthy, non-smoker patients diagnosed with chronic periodontitis were randomly assigned to PRF/DFDBA group or the DFDBA/saline. Clinical [pocket depth (PD), clinical attachment level (CAL) and gingival recession (REC)] and radiographic (bone fill, defect resolution and alveolar crest resorption) measurements were made at baseline and at a 12-month evaluation. Compared with baseline, 12-month results indicated that both treatment modalities resulted in significant changes in all clinical and radiographic parameters. However, the PRP/DFDBA group exhibited statistically significantly greater changes compared with the DFDBA/saline group in PD (4.15 ± 0.84 vs 3.60 ± 0.51 mm), CAL (3.73 ± 0.74 vs 2.61 ± 0.68 mm), REC (0.47 ± 0.56 vs 1.00 ± 0.61 mm), bone fill (3.50 ± 0.67 vs 2.49 ± 0.64 mm) and defect resolution (3.73 ± 0.63 vs 2.75 ± 0.57 mm). Observations indicate that a combination of PRF and DFDBA is more effective than DFDBA with saline for the treatment of infrabony periodontal defects.

  3. Glucose metabolism in cultured trophoblasts from human placenta

    International Nuclear Information System (INIS)

    Moe, A.J.; Farmer, D.R.; Nelson, D.M.; Smith, C.H.

    1990-01-01

    The development of appropriate placental trophoblast isolation and culture techniques enables the study of pathways of glucose utilization by this important cell layer in vitro. Trophoblasts from normal term placentas were isolated and cultured 24 hours and 72 hours in uncoated polystyrene culture tubes or tubes previously coated with a fibrin matrix. Trophoblasts cultured on fibrin are morphologically distinct from those cultured on plastic or other matrices and generally resemble in vivo syncytium. Cells were incubated up to 3 hours with 14 C-labeled glucose and reactions were stopped by addition of perchloric acid. 14 CO 2 production by trophoblasts increased linearly with time however the largest accumulation of label was in organic acids. Trophoblasts cultured in absence of fibrin utilized more glucose and accumulated more 14 C in metabolic products compared to cells cultured on fibrin. Glucose oxidation to CO 2 by the phosphogluconate (PG) pathway was estimated from specific yields of 14 CO 2 from [1- 14 C]-D-glucose and [6- 14 C]-D-glucose. Approximately 6% of glucose oxidation was by the PG pathway when cells were cultured on fibrin compared to approximately 1% by cells cultured in the absence of fibrin. The presence of a fibrin growth matrix appears to modulate the metabolism of glucose by trophoblast from human placenta in vitro

  4. Characterization of Leukocyte-platelet Rich Fibrin, A Novel Biomaterial.

    Science.gov (United States)

    Madurantakam, Parthasarathy; Yoganarasimha, Suyog; Hasan, Fadi K

    2015-09-29

    Autologous platelet concentrates represent promising innovative tools in the field of regenerative medicine and have been extensively used in oral surgery. Unlike platelet rich plasma (PRP) that is a gel or a suspension, Leukocyte-Platelet Rich Fibrin (L-PRF) is a solid 3D fibrin membrane generated chair-side from whole blood containing no anti-coagulant. The membrane has a dense three dimensional fibrin matrix with enriched platelets and abundant growth factors. L-PRF is a popular adjunct in surgeries because of its superior handling characteristics as well as its suturability to the wound bed. The goal of the study is to demonstrate generation as well as provide detailed characterization of relevant properties of L-PRF that underlie its clinical success.

  5. Cartilage oligomeric matrix protein enhances matrix assembly during chondrogenesis of human mesenchymal stem cells.

    Science.gov (United States)

    Haleem-Smith, Hana; Calderon, Raul; Song, Yingjie; Tuan, Rocky S; Chen, Faye H

    2012-04-01

    Cartilage oligomeric matrix protein/thrombospondin-5 (COMP/TSP5) is an abundant cartilage extracellular matrix (ECM) protein that interacts with major cartilage ECM components, including aggrecan and collagens. To test our hypothesis that COMP/TSP5 functions in the assembly of the ECM during cartilage morphogenesis, we have employed mesenchymal stem cell (MSC) chondrogenesis in vitro as a model to examine the effects of COMP over-expression on neo-cartilage formation. Human bone marrow-derived MSCs were transfected with either full-length COMP cDNA or control plasmid, followed by chondrogenic induction in three-dimensional pellet or alginate hydrogel culture. MSC chondrogenesis and ECM production was estimated based on quantitation of sulfated glycosaminoglycan (sGAG) accumulation, immunohistochemistry of the presence and distribution of cartilage ECM proteins, and real-time RT-PCR analyis of mRNA expression of cartilage markers. Our results showed that COMP over-expression resulted in increased total sGAG content during the early phase of MSC chondrogenesis, and increased immuno-detectable levels of aggrecan and collagen type II in the ECM of COMP-transfected pellet and alginate cultures, indicating more abundant cartilaginous matrix. COMP transfection did not significantly increase the transcript levels of the early chondrogenic marker, Sox9, or aggrecan, suggesting that enhancement of MSC cartilage ECM was effected at post-transcriptional levels. These findings strongly suggest that COMP functions in mesenchymal chondrogenesis by enhancing cartilage ECM organization and assembly. The action of COMP is most likely mediated not via direct changes in cartilage matrix gene expression but via interactions of COMP with other cartilage ECM proteins, such as aggrecan and collagens, that result in enhanced assembly and retention.

  6. CARTILAGE OLIGOMERIC MATRIX PROTEIN ENHANCES MATRIX ASSEMBLY DURING CHONDROGENESIS OF HUMAN MESENCHYMAL STEM CELLS

    Science.gov (United States)

    Haleem-Smith, Hana; Calderon, Raul; Song, Yingjie; Tuan, Rocky S.; Chen, Faye H.

    2011-01-01

    Cartilage oligomeric matrix protein/thrombospondin-5 (COMP/TSP5) is an abundant cartilage extracellular matrix (ECM) protein that interacts with major cartilage ECM components, including aggrecan and collagens. To test our hypothesis that COMP/TSP5 functions in the assembly of the ECM during cartilage morphogenesis, we have employed mesenchymal stem cell (MSC) chondrogenesis in vitro as a model to examine the effects of COMP over-expression on neo-cartilage formation. Human bone marrow-derived MSCs were transfected with either full-length COMP cDNA or control plasmid, followed by chondrogenic induction in three-dimensional pellet or alginate-hydrogel culture. MSC chondrogenesis and ECM production was estimated based on quantitation of sulfated glycosaminoglycan (sGAG) accumulation, immunohistochemistry of the presence and distribution of cartilage ECM proteins, and real-time RT-PCR analyis of mRNA expression of cartilage markers. Our results showed that COMP over-expression resulted in increased total sGAG content during the early phase of MSC chondrogenesis, and increased immuno-detectable levels of aggrecan and collagen type II in the ECM of COMP-transfected pellet and alginate cultures, indicating more abundant cartilaginous matrix. COMP transfection did not significantly increase the transcript levels of the early chondrogenic marker, Sox9, or aggrecan, suggesting that enhancement of MSC cartilage ECM was effected at post-transcriptional levels. These findings strongly suggest that COMP functions in mesenchymal chondrogenesis by enhancing cartilage ECM organization and assembly. The action of COMP is most likely mediated not via direct changes in cartilage matrix gene expression but via interactions of COMP with other cartilage ECM proteins, such as aggrecan and collagens, that result in enhanced assembly and retention. PMID:22095699

  7. Effect of dextran on platelet activation by polymerizing fibrin.

    OpenAIRE

    Ts'ao, C. H.; Krajewski, D. V.

    1982-01-01

    Dextran has been shown to alter the mechanical properties and lysability of fibrin. The present study was undertaken to determine whether it would also influence the ability of polymerizing fibrin to activate platelets. Human fibrinogen, with or without the presence of dextran, was incubated with either human thrombin or reptilase at 37 C. Macroscopically evident fibrils first appeared at 7 1/2 to 8 minutes in fibrinogen solution not containing dextran and at 2 1/2 to 3 minutes in solution co...

  8. Experimental closure of gunshot wounds by fibrin glue with antibiotics in pigs

    OpenAIRE

    Đenić Nebojša; Višnjić Milan; Dragović Saša; Bojanić Vladmila; Bojanić Zoran; Đurđević Dragan; Đinđić Boris; Kostov Miloš

    2015-01-01

    Background/Aim. Gunshot wounds caused by the automatic rifle M70AB2 (AK-47) 7.62 mm, after the primary surgical management, were closed with delayed primary suture during the next four to seven days. This period coincides with the fibroblastic phase of wound healing. Fibrin glue is used as a local hemostatic and as a matrix for the local dosed release of antibiotics. Antibiotics addition to fibrin glue resulted in continuous diffusion into the surrounding n...

  9. Fibrin related antigens: assay development, clinical and kinetic studies

    Energy Technology Data Exchange (ETDEWEB)

    Kruskal, J B

    1987-08-01

    This thesis describes an assay which is able to measure and to determine the proportions of fibrin- and fibrinogen-related antigens (FRA) present in clinical samples. No assay exists at present which is capable of distinguishing between fibrin and fibrinogen degradation products concurrently and in a clinical setting. The assay may be used as a tool with which to gain further insight to pathophysiology of disorders characterized by activation of the coagulation and fibrinolytic pathways. This study provides and analysis of the FRA profiles in patients with disorders characterised by possible enhanced fibrinolytic activity. Studies have been undertaken on patients with acute and chronic liver diseases, on patients with the various syndromes of coronary artery disease and on patients with insulin-dependent diabetes mellitus with and without evidence of microvascular disease. Certain observations made it evident that further studies were required in order to explain previously undocumented fibrinolytic abnormalities in certain patient groups. Data obtained from patients with liver disease provided information compatible with the activation of their fibrinolytic pathways. The initial scope of this study was then extended to further investigate the deranged haemostatic mechanisms in patients with severe liver diseases. Kinetic studies were performed which required the development of specific technology to be able to measure certain previously undertermined parameters. Mathematical models describing the rates of fibrin formation and lysis were developed for human studies. Fibrin-derived D-dimer was radiolabelled and its validity as and intravenous tracer and maker of fibrin degradation established.

  10. Extracellular matrix alterations in human corneas with bullous keratopathy

    DEFF Research Database (Denmark)

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

    1996-01-01

    PURPOSE. To uncover abnormalities of extracellular matrix (ECM) distribution in human corneas with pseudophakic and aphakic bullous keratopathy (PBK/ABK). METHODS. Indirect immunofluorescence with antibodies to 27 ECM components was used on frozen sections of 14 normal and 20 PBK/ABK corneas...... in some cases, correlated with decreased visual acuity. In normal central corneas, tenascin was never found. Other major ECM abnormalities in PBK/ABK corneas compared to normals included: discontinuous epithelial BM straining for laminin-1 (alpha 1 beta 1 gamma 1), entactin/nidogen and fibronectin......; accumulation of fibronectin and alpha 1-alpha 2 type IV collagen on the endothelial face of the Descemet's membrane; and abnormal deposition of stromal ECM (tenascin, fibronectin, decorin, types I, III, V, VI, VIII, XII, XIV collagen) and BM components (type IV, collagen, perlecan, bamacan, laminin-1, entactin...

  11. Role of Fibrin Sealants in Liver Surgery

    NARCIS (Netherlands)

    de Boer, Marieke T.; Boonstra, Elizabeth A.; Lisman, Ton; Porte, Robert J.

    2012-01-01

    Background: Fibrin sealants are widely used in liver surgery. The aim of this article is to review the literature on evidence of hemostatic and biliostatic capacities of different fibrin sealants in liver surgery. Methods: In PubMed, a literature search was done with the search terms 'fibrin

  12. Extracellular Matrix Molecular Remodeling in Human Liver Fibrosis Evolution.

    Directory of Open Access Journals (Sweden)

    Andrea Baiocchini

    Full Text Available 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.

  13. A new step toward the artificial ovary: survival and proliferation of isolated murine follicles after autologous transplantation in a fibrin scaffold.

    Science.gov (United States)

    Luyckx, Valérie; Dolmans, Marie-Madeleine; Vanacker, Julie; Legat, Camille; Fortuño Moya, Cristina; Donnez, Jacques; Amorim, Christiani Andrade

    2014-04-01

    To create an artificial ovary to provide an alternative way of restoring fertility in patients who cannot benefit from transplantation of cryopreserved ovarian tissue due to the threat of reintroducing malignant cells. In vivo experimental study. Gynecology research unit in a university hospital. Six-week-old female NMRI mice. Autografting of isolated preantral follicles and ovarian cells (OCs) encapsulated in two fibrin matrices containing low concentrations of fibrinogen (F; mg/mL) and thrombin (T; IU/mL): F12.5/T1 and F25/T4. Follicular density and development, OC survival and proliferation, inflammatory response, and vascularization. After 1 week, the follicle recovery rate ranged from 30.8% (F25/T4) to 31.8% (F12.5/T1). With both fibrin formulations, all follicles were found to be alive or minimally damaged, as demonstrated by terminal deoxynucleotide transferase-mediated dUTP nick-end labeling assay, and at the growing stage (primary, secondary, and antral follicles), confirmed by Ki67 immunostaining. Isolated OCs also survived and proliferated after grafting, as evidenced by <1% apoptotic cells and a high proportion of Ki67-positive cells. Vessels were found in both fibrin formulations, and the global vascular surface area varied from 1.35% (F25/T4) to 1.88% (F12.5/T1). Numerous CD45-positive cells were also observed in both F25/T4 and F12.5/T1 combinations. The present study is the first to show survival and growth of isolated murine ovarian follicles 1 week after autotransplantation of isolated OCs in a fibrin scaffold. The results indicate that fibrin is a promising candidate as a matrix for the construction of an artificial ovary. Xenotransplantation of isolated human follicles and OCs is the necessary next step to validate these findings. Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  14. Differentiation within autologous fibrin scaffolds of porcine dermal cells with the mesenchymal stem cell phenotype

    International Nuclear Information System (INIS)

    Puente, Pilar de la; Ludeña, Dolores; López, Marta; Ramos, Jennifer; Iglesias, Javier

    2013-01-01

    Porcine mesenchymal stem cells (pMSCs) are an attractive source of cells for tissue engineering because their properties are similar to those of human stem cells. pMSCs can be found in different tissues but their dermal origin has not been studied in depth. Additionally, MSCs differentiation in monolayer cultures requires subcultured cells, and these cells are at risk of dedifferentiation when implanting them into living tissue. Following this, we attempted to characterize the MSCs phenotype of porcine dermal cells and to evaluate their cellular proliferation and differentiation in autologous fibrin scaffolds (AFSs). Dermal biopsies and blood samples were obtained from 12 pigs. Dermal cells were characterized by flow cytometry. Frozen autologous plasma was used to prepare AFSs. pMSC differentiation was studied in standard structures (monolayers and pellets) and in AFSs. The pMSCs expressed the CD90 and CD29 markers of the mesenchymal lineage. AFSs afforded adipogenic, osteogenic and chondrogenic differentiation. The porcine dermis can be proposed to be a good source of MSCs with adequate proliferative capacity and a suitable expression of markers. The pMSCs also showed optimal proliferation and differentiation in AFSs, such that these might serve as a promising autologous and implantable material for use in tissue engineering. -- Highlights: ► Low fibrinogen concentration provides a suitable matrix for cell migration and differentiation. ► Autologous fibrin scaffolds is a promising technique in tissue engineering. ► Dermal cells are an easily accessible mesenchymal stem cell source. ► Fibrin scaffolds afforded adipogenic, osteogenic and chondrogenic differentiation.

  15. Fibrin gel as a scaffold for skin substitute – production and clinical experience.

    Science.gov (United States)

    Kljenak, Antun; Tominac Trcin, Mirna; Bujić, Marina; Dolenec, Tamara; Jevak, Martina; Mršić, Gordan; Zmiš, Gordana; Barčot, Zoran; Muljačić, Ante; Popović, Maja

    2016-06-01

    The purpose of this study was to create a fibrin-based human skin substitute in vitro with epidermal and dermal component and to assess its healing potential in deep partial and full thickness burns. Fibrin scaffolds were prepared from commercial fibrin glue kits. Human fibroblasts were cultured in fibrin gel. Human keratinocytes were seeded on the top of the gel. Viability of cells was determined fluorimetrically. Scanning electron microscope and immunocytochemistry analysis of cultured cells were performed. After hydrosurgical preparation of deep burn necrotic tissue, wound bed was prepared for skin substitutes. Progress of healing was documented using visual estimation and photos. Scanning electron microscope images showed good cell attachment and colony spreading of keratinocytes and fibroblasts on fibrin scaff old. Immunofluorescent staining of cell cultures on fibrin scaffold showed expression of vimentin, a marker of fibroblast cells, cytokeratin 19, a marker of epithelial stem cells, as well as involucrin, a marker of differentiated keratinocytes. Clinical results clearly showed that appearance of the skin did not differ significantly from the areas of transplanted skin using split-thickness skin graft techniques. In conclusion, using these fibrin-cultured autografts on massive full-thickness burn resulted in good healing.

  16. SPECT imaging of fibrin using fibrin-binding peptides

    NARCIS (Netherlands)

    Starmans, L.W.E.; Duijnhoven, van S.M.J.; Rossin, R.; Aime, S.; Daemen, M.J.A.P.; Nicolay, K.; Grüll, H.

    2013-01-01

    Noninvasive detection of fibrin in vivo using diagnostic imaging modalities may improve clinical decision-making on possible therapeutic options in atherosclerosis, cancer and thrombus-related pathologies such as pulmonary embolism and deep venous thrombosis. The aim of this study was to assess the

  17. Fibrin accumulation secondary to loss of plasmin-mediated fibrinolysis drives inflammatory osteoporosis in mice.

    Science.gov (United States)

    Cole, Heather A; Ohba, Tetsuro; Nyman, Jeffry S; Hirotaka, Haro; Cates, Justin M M; Flick, Matthew J; Degen, Jay L; Schoenecker, Jonathan G

    2014-08-01

    Osteoporosis is a skeletal disorder characterized by low bone mass and increased bone fragility associated with aging, menopause, smoking, obesity, or diabetes. Persistent inflammation has been identified as an instigating factor in progressive bone loss. In addition to the role of fibrin in coagulation, inordinate fibrin deposition within a tissue matrix results in increased local inflammation. Given that fibrin accumulation is a hallmark of osteoporosis-related comorbidities, we undertook this study to test the hypothesis that persistent fibrin deposition causes inflammatory osteoporosis. Multiple imaging modalities, bone integrity metrics, and histologic analyses were employed to evaluate skeletal derangements in relation to fibrin deposition, circulating fibrinogen levels, and systemic markers of inflammation in mice that were plasminogen deficient and in plasminogen-deficient mice that were concomitantly either fibrinogen deficient or carrying a mutant form of fibrinogen lacking the αM β2 binding motif. Mice generated with a genetic deficit in the key fibrinolytic protease, plasmin, uniformly developed severe osteoporosis. Furthermore, the development of osteoporosis was fibrin(ogen) dependent, and the derangements in the bone remodeling unit were mechanistically tied to fibrin(ogen)-mediated activation of osteoclasts via activation of the leukocyte integrin receptor αM β2 on monocytes and secondary stimulation of osteoblasts by RANKL. Notably, the genetic elimination of fibrin(ogen) or the expression of a mutant form of fibrinogen retaining clotting function but lacking the αM β2 binding motif prevented the degenerative skeletal phenotypes, resulting in normal local and systemic cytokine levels. Taken together, these data reveal for the first time that fibrin promotes inflammation-driven systemic osteoporosis, which suggests a novel association between hemostasis, inflammation, and bone biology. Copyright © 2014 by the American College of Rheumatology.

  18. Platelet-rich fibrin versus albumin in surgical wound repair: a randomized trial with paired design

    DEFF Research Database (Denmark)

    Danielsen, Patricia L; Ågren, Sven Per Magnus; Jørgensen, Lars Nannestad

    2010-01-01

    To study the effects of autologous platelet-rich fibrin (PRF) versus human albumin on incisional wound breaking strength and subcutaneous collagen deposition in patients undergoing laparoscopic cholecystectomy in a randomized trial.......To study the effects of autologous platelet-rich fibrin (PRF) versus human albumin on incisional wound breaking strength and subcutaneous collagen deposition in patients undergoing laparoscopic cholecystectomy in a randomized trial....

  19. Proliferation and differentiation of stem cells in contact with eluate from fibrin-rich plasma membrane

    Directory of Open Access Journals (Sweden)

    Fernanda Gimenez de Souza

    Full Text Available ABSTRACT Objective: To evaluate the ability of the eluate from fibrin-rich plasma (FRP membrane to induce proliferation and differentiation of isolated human adipose-derived stem cells (ASCs into chondrocytes. Method: FRP membranes were obtained by centrifugation of peripheral blood from two healthy donors, cut, and maintained in culture plate wells for 48 h to prepare the fibrin eluate. The SCATh were isolated from adipose tissue by collagenase digestion solution, and expanded in vitro. Cells were expanded and treated with DMEM-F12 culture, a commercial media for chondrogenic differentiation, and eluate from FRP membrane for three days, and labeled with BrdU for quantitative assessment of cell proliferation using the High-Content Operetta® imaging system. For the chondrogenic differentiation assay, the SCATh were grown in micromass for 21 days and stained with toluidine blue and aggrecan for qualitative evaluation by light microscopy. The statistical analysis was performed using ANOVA and Tukey's test. Results: There was a greater proliferation of cells treated with the eluate from FRP membrane compared to the other two treatments, where the ANOVA test showed significance (p < 0.001. The differentiation into chondrocytes was visualized by the presence of mucopolysaccharide in the matrix of the cells marked in blue toluidine and aggrecan. Conclusions: Treatment with eluate from FRP membrane stimulated cell proliferation and induced differentiation of the stem cells into chondrocytes, suggesting a potential application of FRP membranes in hyaline cartilage regeneration therapies.

  20. Concentration independent modulation of local micromechanics in a fibrin gel.

    Directory of Open Access Journals (Sweden)

    Maxwell A Kotlarchyk

    Full Text Available Methods for tuning extracellular matrix (ECM mechanics in 3D cell culture that rely on increasing the concentration of either protein or cross-linking molecules fail to control important parameters such as pore size, ligand density, and molecular diffusivity. Alternatively, ECM stiffness can be modulated independently from protein concentration by mechanically loading the ECM. We have developed a novel device for generating stiffness gradients in naturally derived ECMs, where stiffness is tuned by inducing strain, while local mechanical properties are directly determined by laser tweezers based active microrheology (AMR. Hydrogel substrates polymerized within 35 mm diameter Petri dishes are strained non-uniformly by the precise rotation of an embedded cylindrical post, and exhibit a position-dependent stiffness with little to no modulation of local mesh geometry. Here we present the device in the context of fibrin hydrogels. First AMR is used to directly measure local micromechanics in unstrained hydrogels of increasing fibrin concentration. Changes in stiffness are then mapped within our device, where fibrin concentration is held constant. Fluorescence confocal imaging and orbital particle tracking are used to quantify structural changes in fibrin on the micro and nano levels respectively. The micromechanical strain stiffening measured by microrheology is not accompanied by ECM microstructural changes under our applied loads, as measured by confocal microscopy. However, super-resolution orbital tracking reveals nanostructural straightening, lengthening, and reduced movement of fibrin fibers. Furthermore, we show that aortic smooth muscle cells cultured within our device are morphologically sensitive to the induced mechanical gradient. Our results demonstrate a powerful cell culture tool that can be used in the study of mechanical effects on cellular physiology in naturally derived 3D ECM tissues.

  1. Adipose-Derived-Stem-Cell-Seeded Fibrin Matrices for Periodontal Ligament Engineering: The Need for Dynamic Strain

    NARCIS (Netherlands)

    de Jong, Thijs; Oostendorp, Corien; Bakker, Astrid D.; van Kuppevelt, Toin H.; Smit, Theo H.

    2017-01-01

    Introduction: The periodontal ligament (PDL) connects the tooth to the alveolar bone. For PDL regeneration after tissue damage, we propose human adipose-derived stem cells (hASCs) embedded in fibrin. We showed previously that hASCs in fibrin extensively produce collagen, but in a non-functional,

  2. Effects of the incorporation of ε-aminocaproic acid/chitosan particles to fibrin on cementoblast differentiation and cementum regeneration.

    Science.gov (United States)

    Park, Chan Ho; Oh, Joung-Hwan; Jung, Hong-Moon; Choi, Yoonnyoung; Rahman, Saeed Ur; Kim, Sungtae; Kim, Tae-Il; Shin, Hong-In; Lee, Yun-Sil; Yu, Frank H; Baek, Jeong-Hwa; Ryoo, Hyun-Mo; Woo, Kyung Mi

    2017-10-01

    Cementum formation on the exposed tooth-root surface is a critical process in periodontal regeneration. Although various therapeutic approaches have been developed, regeneration of integrated and functional periodontal complexes is still wanting. Here, we found that the OCCM30 cementoblasts cultured on fibrin matrix express substantial levels of matrix proteinases, leading to the degradation of fibrin and the apoptosis of OCCM30 cells, which was reversed upon treatment with a proteinase inhibitor, ε-aminocaproic acid (ACA). Based on these findings, ACA-releasing chitosan particles (ACP) were fabricated and ACP-incorporated fibrin (fibrin-ACP) promoted the differentiation of cementoblasts in vitro, as confirmed by bio-mineralization and expressions of molecules associated with mineralization. In a periodontal defect model of beagle dogs, fibrin-ACP resulted in substantial cementum formation on the exposed root dentin in vivo, compared to fibrin-only and enamel matrix derivative (EMD) which is used clinically for periodontal regeneration. Remarkably, the fibrin-ACP developed structural integrations of the cementum-periodontal ligament-bone complex by the Sharpey's fiber insertion. In addition, fibrin-ACP promoted alveolar bone regeneration through increased bone volume of tooth roof-of-furcation defects and root coverage. Therefore, fibrin-ACP can promote cementogenesis and osteogenesis by controlling biodegradability of fibrin, implicating the feasibility of its therapeutic use to improve periodontal regeneration. Cementum, the mineralized layer on root dentin surfaces, functions to anchor fibrous connective tissues on tooth-root surfaces with the collagenous Sharpey's fibers integration, of which are essential for periodontal functioning restoration in the complex. Through the cementum-responsible fiber insertions on tooth-root surfaces, PDLs transmit various mechanical responses to periodontal complexes against masticatory/occlusal stimulations to support teeth

  3. Effectiveness of Fibrin Glue in Adherence of Skin Graft.

    Science.gov (United States)

    Reddy, Konda Sireesha; Chittoria, Ravi Kumar; Babu, Preethitha; Marimuthu, Senthil Kumaran; Kumar, Sudhanva Hemanth; Subbarayan, Elan Kumar; Chavan, Vinayak; Mohapatra, Devi Prasad; Sivakumar, Dinesh Kumar; Friji, M T

    2017-01-01

    Graft fixation is important for graft take. Fibrin glue has been proposed as an ideal material, because of its human origin and it provides firm adhesion in seconds or minutes. To evaluate the efficiency of fibrin glue, in increasing the take of skin graft. Assessment includes surgical time taken for graft fixation, haematoma/seroma formation, engraftment and wound closure by day 14. The study is an observational prospective study conducted in the Department of Plastic Surgery, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry, from January 2016 to June 2016. Sixteen patients who underwent split skin grafting were assessed during the study period. Fibrin glue was used on the recipient bed before grafting. Better haemostasis and graft adhesion, with a significant reduction of surgical time, were noted. The safety profile of fibrin glue was excellent as indicated by the lack of any related serious adverse experiences. These findings demonstrate that it is safe and effective for attachment of skin grafts, with outcomes at least as good as conventional methods.

  4. In vivo extracellular matrix protein expression by human periodontal ...

    African Journals Online (AJOL)

    ONOS

    2010-08-23

    Aug 23, 2010 ... Extracellular matrix proteins (ECM) are described as molecular regulators of these events. ..... zation and adhesive interaction of cells (Yamada, 1983). .... periodontal ligament fibroblasts after simulation of orthodontic force.

  5. Fibrin nanostructures for biomedical applications

    Czech Academy of Sciences Publication Activity Database

    Riedelová-Reicheltová, Zuzana; Brynda, Eduard; Riedel, Tomáš

    2016-01-01

    Roč. 65, Suppl. 2 (2016), S263-S272 ISSN 0862-8408 R&D Projects: GA MŠk(CZ) LQ1604 Institutional support: RVO:61389013 Keywords : fibrinogen * fibrin-bound thrombin * nanostructures Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.461, year: 2016 http://www.biomed.cas.cz/physiolres/pdf/65%20Suppl%202/65_S263.pdf

  6. Three-dimensional bone tissue substitute based on a human mesenchymal stem cell culture on a nanofiber carrier and inorganic matrix

    Directory of Open Access Journals (Sweden)

    Martin Krbec

    2016-01-01

    Full Text Available The aim was to construct a composite structure for bone tissue substitute on the basis of a degradable composite of an organic nanofiber carrier and an inorganic matrix in 3D, and to achieve subsequent colonisation by differentiated human mesenchymal stem cells (hMSC towards osteocytes. We developed an active bone tissue substitute using nanofiber technology for a polycaprolactone (PCL scaffold with the addition of hydroxyapatite and the colonisation of both components with hMSC with the ability of differentiation towards osteocytes. The constructed composition included the components necessary for bone healing (inorganic and cellular and it also forms a spatially-oriented 3D structure. We used polycaprolactone Mw 70,000 with electrostatic spinning for the formation of nanofibers using a modified NanospiderTM method. For the inorganic component we used orthophosphate-calcium silicate with a crystal size of 1-2 mm which the nanofiber membrane was coated with. Both components were connected together with a tissue adhesive based of fibrin glue. Cultivated hMSC cells at a concentration of 1.2 × 104/cm2 were multiplied in vitro and then cultivated in the expansion medium. HMSC overgrew both the PCL membrane and the Si-CaP crystals. After colonisation with cultivated cells, this composite 3D structure can serve as a three-dimensional bone tissue replacement.

  7. HGF potentiates extracellular matrix-driven migration of human myoblasts: involvement of matrix metalloproteinases and MAPK/ERK pathway.

    Science.gov (United States)

    González, Mariela Natacha; de Mello, Wallace; Butler-Browne, Gillian S; Silva-Barbosa, Suse Dayse; Mouly, Vincent; Savino, Wilson; Riederer, Ingo

    2017-10-10

    The hepatocyte growth factor (HGF) is required for the activation of muscle progenitor cells called satellite cells (SC), plays a role in the migration of proliferating SC (myoblasts), and is present as a soluble factor during muscle regeneration, along with extracellular matrix (ECM) molecules. In this study, we aimed at determining whether HGF is able to interact with ECM proteins, particularly laminin 111 and fibronectin, and to modulate human myoblast migration. We evaluated the expression of the HGF-receptor c-Met, laminin, and fibronectin receptors by immunoblotting, flow cytometry, or immunofluorescence and used Transwell assays to analyze myoblast migration on laminin 111 and fibronectin in the absence or presence of HGF. Zymography was used to check whether HGF could modulate the production of matrix metalloproteinases by human myoblasts, and the activation of MAPK/ERK pathways was evaluated by immunoblotting. We demonstrated that human myoblasts express c-Met, together with laminin and fibronectin receptors. We observed that human laminin 111 and fibronectin have a chemotactic effect on myoblast migration, and this was synergistically increased when low doses of HGF were added. We detected an increase in MMP-2 activity in myoblasts treated with HGF. Conversely, MMP-2 inhibition decreased the HGF-associated stimulation of cell migration triggered by laminin or fibronectin. HGF treatment also induced in human myoblasts activation of MAPK/ERK pathways, whose specific inhibition decreased the HGF-associated stimulus of cell migration triggered by laminin 111 or fibronectin. We demonstrate that HGF induces ERK phosphorylation and MMP production, thus stimulating human myoblast migration on ECM molecules. Conceptually, these data state that the mechanisms involved in the migration of human myoblasts comprise both soluble and insoluble moieties. This should be taken into account to optimize the design of therapeutic cell transplantation strategies by improving

  8. Three-dimensional structure and cytokine distribution of platelet-rich fibrin.

    Science.gov (United States)

    Bai, Meng-Yi; Wang, Ching-Wei; Wang, Jyun-Yi; Lin, Ming-Fang; Chan, Wing P

    2017-02-01

    Previous reports have revealed that several cytokines (including platelet-derived growth factor-BB, transforming growth factors-β1 and insulin-like growth factor-1) can enhance the rate of bone formation and synthesis of extracellular matrix in orthopaedics or periodontology. This study aimed to determine the concentration of cytokines within platelet-rich fibrin microstructures and investigate whether there are differences in the different portions of platelet-rich fibrin, which has implications for proper clinical use of platelet-rich fibrin gel. Whole blood was obtained from six New Zealand rabbits (male, 7 to 39 weeks old, weight 2.7-4 kg); it was then centrifuged for preparation of platelet-rich fibrin gels and harvest of plasma. The resultant platelet-rich fibrin gels were used for cytokine determination, histological analyses and scanning electron microscopy. All plasmas obtained were subject to the same cytokine determination assays for the purpose of comparison. Cytokines platelet-derived growth factor-BB and transforming growth factor-β1 formed concentration gradients from high at the red blood cell end of the platelet-rich fibrin gel (p=1.88×10-5) to low at the plasma end (p=0.19). Insulin-like growth factor-1 concentrations were similar at the red blood cell and plasma ends. The porosities of the platelet-rich fibrin samples taken in sequence from the red blood cell end to the plasma end were 6.5% ± 4.9%, 24.8% ± 7.5%, 30.3% ± 8.5%, 41.4% ± 12.3%, and 40.3% ± 11.7%, respectively, showing a gradual decrease in the compactness of the platelet-rich fibrin network. Cytokine concentrations are positively associated with platelet-rich fibrin microstructure and portion in a rabbit model. As platelet-rich fibrin is the main entity currently used in regenerative medicine, assessing cytokine concentration and the most valuable portion of PRF gels is essential and recommended to all physicians.

  9. Three-dimensional structure and cytokine distribution of platelet-rich fibrin

    Directory of Open Access Journals (Sweden)

    Meng-Yi Bai

    Full Text Available OBJECTIVES: Previous reports have revealed that several cytokines (including platelet-derived growth factor-BB, transforming growth factors-β1 and insulin-like growth factor-1 can enhance the rate of bone formation and synthesis of extracellular matrix in orthopaedics or periodontology. This study aimed to determine the concentration of cytokines within platelet-rich fibrin microstructures and investigate whether there are differences in the different portions of platelet-rich fibrin, which has implications for proper clinical use of platelet-rich fibrin gel. METHODS: Whole blood was obtained from six New Zealand rabbits (male, 7 to 39 weeks old, weight 2.7-4 kg; it was then centrifuged for preparation of platelet-rich fibrin gels and harvest of plasma. The resultant platelet-rich fibrin gels were used for cytokine determination, histological analyses and scanning electron microscopy. All plasmas obtained were subject to the same cytokine determination assays for the purpose of comparison. RESULTS: Cytokines platelet-derived growth factor-BB and transforming growth factor-β1 formed concentration gradients from high at the red blood cell end of the platelet-rich fibrin gel (p=1.88×10-5 to low at the plasma end (p=0.19. Insulin-like growth factor-1 concentrations were similar at the red blood cell and plasma ends. The porosities of the platelet-rich fibrin samples taken in sequence from the red blood cell end to the plasma end were 6.5% ± 4.9%, 24.8% ± 7.5%, 30.3% ± 8.5%, 41.4% ± 12.3%, and 40.3% ± 11.7%, respectively, showing a gradual decrease in the compactness of the platelet-rich fibrin network. CONCLUSION: Cytokine concentrations are positively associated with platelet-rich fibrin microstructure and portion in a rabbit model. As platelet-rich fibrin is the main entity currently used in regenerative medicine, assessing cytokine concentration and the most valuable portion of PRF gels is essential and recommended to all physicians.

  10. A novel platelet concentrate: titanium-prepared platelet-rich fibrin.

    Science.gov (United States)

    Tunalı, Mustafa; Özdemir, Hakan; Küçükodacı, Zafer; Akman, Serhan; Yaprak, Emre; Toker, Hülya; Fıratlı, Erhan

    2014-01-01

    We developed a new product called titanium-prepared platelet-rich fibrin (T-PRF). The T-PRF method is based on the hypothesis that titanium may be more effective in activating platelets than the silica activators used with glass tubes in Chouckroun's leukocyte- and platelet-rich fibrin (L-PRF) method. In this study, we aimed to define the structural characteristics of T-PRF and compare it with L-PRF. Blood samples were collected from 10 healthy male volunteers. The blood samples were drawn using a syringe. Nine milliliters was transferred to a dry glass tube, and 9 mL was transferred to a titanium tube. Half of each clot (i.e., the blood that was clotted using T-PRF or L-PRF) was processed with a scanning electron microscope (SEM). The other half of each clot was processed for fluorescence microscopy analysis and light microscopy analysis. The T-PRF samples seemed to have a highly organized network with continuous integrity compared to the other L-PRF samples. Histomorphometric analysis showed that T-PRF fibrin network covers larger area than L-PRF fibrin network; also fibrin seemed thicker in the T-PRF samples. This is the first human study to define T-PRF as an autogenous leukocyte- and platelet-rich fibrin product. The platelet activation by titanium seems to offer some high characteristics to T-PRF.

  11. A Novel Platelet Concentrate: Titanium-Prepared Platelet-Rich Fibrin

    Directory of Open Access Journals (Sweden)

    Mustafa Tunalı

    2014-01-01

    Full Text Available We developed a new product called titanium-prepared platelet-rich fibrin (T-PRF. The T-PRF method is based on the hypothesis that titanium may be more effective in activating platelets than the silica activators used with glass tubes in Chouckroun’s leukocyte- and platelet-rich fibrin (L-PRF method. In this study, we aimed to define the structural characteristics of T-PRF and compare it with L-PRF. Blood samples were collected from 10 healthy male volunteers. The blood samples were drawn using a syringe. Nine milliliters was transferred to a dry glass tube, and 9 mL was transferred to a titanium tube. Half of each clot (i.e., the blood that was clotted using T-PRF or L-PRF was processed with a scanning electron microscope (SEM. The other half of each clot was processed for fluorescence microscopy analysis and light microscopy analysis. The T-PRF samples seemed to have a highly organized network with continuous integrity compared to the other L-PRF samples. Histomorphometric analysis showed that T-PRF fibrin network covers larger area than L-PRF fibrin network; also fibrin seemed thicker in the T-PRF samples. This is the first human study to define T-PRF as an autogenous leukocyte- and platelet-rich fibrin product. The platelet activation by titanium seems to offer some high characteristics to T-PRF.

  12. A safe and efficient method to retrieve mesenchymal stem cells from three-dimensional fibrin gels.

    Science.gov (United States)

    Carrion, Bita; Janson, Isaac A; Kong, Yen P; Putnam, Andrew J

    2014-03-01

    Mesenchymal stem cells (MSCs) display multipotent characteristics that make them ideal for potential therapeutic applications. MSCs are typically cultured as monolayers on tissue culture plastic, but there is increasing evidence suggesting that they may lose their multipotency over time in vitro and eventually cease to retain any resemblance to in vivo resident MSCs. Three-dimensional (3D) culture systems that more closely recapitulate the physiological environment of MSCs and other cell types are increasingly explored for their capacity to support and maintain the cell phenotypes. In much of our own work, we have utilized fibrin, a natural protein-based material that serves as the provisional extracellular matrix during wound healing. Fibrin has proven to be useful in numerous tissue engineering applications and has been used clinically as a hemostatic material. Its rapid self-assembly driven by thrombin-mediated alteration of fibrinogen makes fibrin an attractive 3D substrate, in which cells can adhere, spread, proliferate, and undergo complex morphogenetic programs. However, there is a significant need for simple cost-effective methods to safely retrieve cells encapsulated within fibrin hydrogels to perform additional analyses or use the cells for therapy. Here, we present a safe and efficient protocol for the isolation of MSCs from 3D fibrin gels. The key ingredient of our successful extraction method is nattokinase, a serine protease of the subtilisin family that has a strong fibrinolytic activity. Our data show that MSCs recovered from 3D fibrin gels using nattokinase are not only viable but also retain their proliferative and multilineage potentials. Demonstrated for MSCs, this method can be readily adapted to retrieve any other cell type from 3D fibrin gel constructs for various applications, including expansion, bioassays, and in vivo implantation.

  13. Acute fibrinous and organising pneumonia.

    Science.gov (United States)

    Gonçalves, João Rocha; Marques, Ricardo; Serra, Paula; Cardoso, Leila

    2017-09-07

    Acute fibrinous and organising pneumonia (AFOP) is a rare histological pattern of interstitial lung disease. The authors describe a 60-year-old woman admitted to the hospital for sustained fever, presenting with an alveolar opacity on chest X-ray, with the presumed diagnosis of community-acquired pneumonia and the onset of antibiotics. Since serological results suggested that Legionella pneumophila was the infectious agent, she was discharged on levofloxacin. A week later, she was again admitted with fever. CT scan showed opacities with crescentic morphology and a central ground-glass area suggestive of cryptogenic organising pneumonia. Microbiological, serological and autoimmunity tests were negative. She underwent surgical lung biopsy that revealed inflammatory infiltrate, macrophage desquamation, fibroblasts proliferation and fibrin deposition in the alveolar spaces, consistent with AFOP. She started corticotherapy with good response. Disease relapsed after prednisolone discontinuation, 10 months later. Currently, the patient is on prednisolone 5 mg/day without clinical and radiological recurrence. © BMJ Publishing Group Ltd (unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  14. Bee venom induces apoptosis and suppresses matrix metaloprotease-2 expression in human glioblastoma cells

    Directory of Open Access Journals (Sweden)

    Mohsen Sisakht

    Full Text Available Abstract Glioblastoma is the most common malignant brain tumor representing with poor prognosis, therapy resistance and high metastasis rate. Increased expression and activity of matrix metalloproteinase-2, a member of matrix metalloproteinase family proteins, has been reported in many cancers including glioblastoma. Inhibition of matrix metalloproteinase-2 expression has resulted in reduced aggression of glioblastoma tumors in several reports. In the present study, we evaluated effect of bee venom on expression and activity of matrix metalloproteinase-2 as well as potential toxicity and apoptogenic properties of bee venom on glioblastoma cells. Human A172 glioblastoma cells were treated with increasing concentrations of bee venom. Then, cell viability, apoptosis, matrix metalloproteinase-2 expression, and matrix metalloproteinase-2 activity were measured using MMT assay, propidium iodide staining, real time-PCR, and zymography, respectively. The IC50 value of bee venom was 28.5 µg/ml in which it leads to decrease of cell viability and induction of apoptosis. Incubation with bee venom also decreased the expression of matrix metalloproteinase-2 in this cell line (p < 0.05. In zymography, there was a reverse correlation between bee venom concentration and total matrix metalloproteinase-2 activity. Induction of apoptosis as well as inhibition of matrix metalloproteinase-2 activity and expression can be suggested as molecular mechanisms involved in cytotoxic and antimetastatic effects of bee venom against glioblastoma cells.

  15. First step in developing a 3D biodegradable fibrin scaffold for an artificial ovary.

    Science.gov (United States)

    Luyckx, Valérie; Dolmans, Marie-Madeleine; Vanacker, Julie; Scalercio, Sarah R; Donnez, Jacques; Amorim, Christiani A

    2013-11-25

    Although transplantation of cryopreserved ovarian tissue is a promising approach to restore fertility in cancer patients, it is not advisable for women at risk of ovarian involvement due to the threat of reintroducing malignant cells. The aim of this study was therefore to find an alternative for these patients by development of an artificial ovary. For construction of the artificial ovary matrix, we used a central composite design to investigate nine combinations of fibrinogen (mg/ml) and thrombin (IU/mL) (F/T): F1/T4, F12.5/T1, F12.5/T20, F25/T0.1, F25/T4, F25/T500, F50/T1, F50/T20 and F100/T4. From the first qualitative analyses (handling and matrix size), five combinations (F12.5/T1, F25/T4, F50/T20, F50/T1 and F100/T4) yielded positive results. They were further evaluated in order to assess fibrin matrix degradation and homogeneous cell encapsulation (density), survival and proliferation (Ki67), and atresia (TUNEL) before and after 7 days of in vitro culture. To determine the best compromise between maximizing the dynamic density (Y1) and minimizing the apoptosis rate (Y2), we used the desirability function approach. Two combinations (F12.5/T1 and F25/T4) showed greater distribution of cells before in vitro culture, reproducible degradation of the fibrin network and adequate support for isolated human ovarian stromal cells, with a high proportion of Ki67-positive cells. SEM analysis revealed a network of fibers with regular pores and healthy stromal cells after in vitro culture with both F/T combinations. This study reports two optimal F/T combinations that allow survival and proliferation of isolated human ovarian cells. Further studies are required to determine if such a scaffold will also be a suitable environment for isolated ovarian follicles.

  16. Matrix and Tensor Completion on a Human Activity Recognition Framework.

    Science.gov (United States)

    Savvaki, Sofia; Tsagkatakis, Grigorios; Panousopoulou, Athanasia; Tsakalides, Panagiotis

    2017-11-01

    Sensor-based activity recognition is encountered in innumerable applications of the arena of pervasive healthcare and plays a crucial role in biomedical research. Nonetheless, the frequent situation of unobserved measurements impairs the ability of machine learning algorithms to efficiently extract context from raw streams of data. In this paper, we study the problem of accurate estimation of missing multimodal inertial data and we propose a classification framework that considers the reconstruction of subsampled data during the test phase. We introduce the concept of forming the available data streams into low-rank two-dimensional (2-D) and 3-D Hankel structures, and we exploit data redundancies using sophisticated imputation techniques, namely matrix and tensor completion. Moreover, we examine the impact of reconstruction on the classification performance by experimenting with several state-of-the-art classifiers. The system is evaluated with respect to different data structuring scenarios, the volume of data available for reconstruction, and various levels of missing values per device. Finally, the tradeoff between subsampling accuracy and energy conservation in wearable platforms is examined. Our analysis relies on two public datasets containing inertial data, which extend to numerous activities, multiple sensing parameters, and body locations. The results highlight that robust classification accuracy can be achieved through recovery, even for extremely subsampled data streams.

  17. Utilizing the Human, Machine, and Environment Matrix in investigations

    International Nuclear Information System (INIS)

    Curry, David; McKinney, John M.

    2006-01-01

    'How did we get into this situation?' How many times has this question been asked at the outset of an investigation, or more importantly, at the completion of an investigation? If the answer is not readily and thoroughly apparent, the investigation is not complete. Subsequently, those who will have the responsibility for correction of the conditions leading to the incident will not have all the information necessary to properly complete their task. For many years, in many writings, the Human/Machine interaction and its impact on process design has been discussed. The same impact should be examined when performing incident investigations. Consideration of the interaction of human and machine along with the environment in which they are used has long been recommended by the National Safety Council, in both design and investigation

  18. A review of bioceramics and fibrin sealant.

    Science.gov (United States)

    Le Guéhennec, L; Layrolle, P; Daculsi, G

    2004-09-13

    This review focuses on bone substitute composites made by mixing ceramic biomaterials with fibrin sealants. Different biomaterials such as coral, bone-derived materials, bioactive glass ceramics, and synthetic calcium phosphate have been mixed with fibrin sealant, resulting in a combination of the biological properties of the two components. This type of association has not produced identical results in all studies. In the past for some, the addition of fibrin sealant to the biomaterial failed to produce any significant, positive effect on osteointegration, whereas others found a positive impact on bone colonization. Despite the negative biological effects reported previously, bioceramic-fibrin composites have been widely used in various types of bone surgery because they are easy to manipulate. In particular, the intra-operative preparation of these composites makes it possible to add bone growth factors or autologous osteoprogenitor cells prior to bone reconstruction. The bone growth factors and autologous osteoprogenitor cells associated with the bioceramic-fibrin composites should provide surgeons with tissue engineered grafts with enhanced osteointegrative properties. This review discusses both the advantages and disadvantages, as well as the future perspectives, of using bioceramic-fibrin composites in various clinical indications.

  19. A Review of Bioceramics and Fibrin Sealant

    Directory of Open Access Journals (Sweden)

    Le Guéhennec L.

    2004-09-01

    Full Text Available This review focuses on bone substitute composites made by mixing ceramic biomaterials with fibrin sealants. Different biomaterials such as coral, bone-derived materials, bioactive glass ceramics, and synthetic calcium phosphate have been mixed with fibrin sealant, resulting in a combination of the biological properties of the two components. This type of association has not produced identical results in all studies. In the past for some, the addition of fibrin sealant to the biomaterial failed to produce any significant, positive effect on osteointegration, whereas others found a positive impact on bone colonization. Despite the negative biological effects reported previously, bioceramic-fibrin composites have been widely used in various types of bone surgery because they are easy to manipulate. In particular, the intra-operative preparation of these composites makes it possible to add bone growth factors or autologous osteoprogenitor cells prior to bone reconstruction. The bone growth factors and autologous osteoprogenitor cells associated with the bioceramic-fibrin composites should provide surgeons with tissue engineered grafts with enhanced osteointegrative properties. This review discusses both the advantages and disadvantages, as well as the future perspectives, of using bioceramic-fibrin composites in various clinical indications.

  20. Coverage of Megaprosthesis with Human Acellular Dermal Matrix after Ewing's Sarcoma Resection: A Case Report

    Directory of Open Access Journals (Sweden)

    Robert M. Whitfield

    2011-01-01

    Full Text Available A 23-year-old female with Ewing's Sarcoma underwent tibial resection and skeletal reconstruction using proximal tibial allograft prosthetic reconstruction with distal femur endoprosthetic reconstruction and rotating hinge. Human acellular dermal matrix, (Alloderm, LifeCell, Branchburg, NJ, USA, was used to wrap the skeletal reconstruction. Soft tissue reconstruction was completed with a rotational gastrocnemius muscle flap and skin graft. Despite prolonged immobilization, the patient quickly regained full range of motion of her skeletal reconstruction. Synthetic mesh, tapes and tubes are used to perform capsule reconstruction of megaprosthesis. This paper describes the role of human acellular dermal matrix in capsule reconstruction around a megaprosthesis.

  1. Determination of fibrin glue with antibiotics on collagen production in colon anastomosis

    Directory of Open Access Journals (Sweden)

    Stanojković Zoran

    2008-01-01

    Full Text Available Background/Aim. Fibrin glue is used as a matrix for local application of antibiotics. The aim of this study was to determine whether application of fibrin glue in combination with antibiotics can strengthen collagen production, prevent dehiscence of colon anastomoses due to infection, and reduce frequency of mortality and morbidity comparing to the control group and the group with fibrin glue application. Methods. The adult male Wistar rats divided into three groups were used in the experiment. The group 1 was the control one (after partial colon resection, colonic anastomoses performed were not treated, while to the group 2 and the group 3 were applied fibrin glue and fibrin glue with antibiotics (clindamycin and ceftriaxon on the site of anastomoses, respectively. Quality of colonic anastomoses were estimated by means of determination of collagen (L-hydroxyproline amount in the collon wall with anastomoses and histological analysis of this colon segment using light and electronic microscope on the days 5, 7 and 13 postoperatively. Results. The highest morbidity rate was registered in the group 1 (30%, then in the group 2 (13.3% and the lowest one in the group 3 (3.33%; p < 0,05 vs group 1. Mortality rate was significantly higher in the group 1 than in the group 3 (20% and 0%, respectively; p < 0,05. In the postoperative course, the highest concentrations of collagen in the colon wall on the site of anastomoses, which was confirmed by both light and electronic microscopy, were found in the group 3. Conclusion. The application of fibrin glue with antibiotics on colon anastomoses reduces the number of dehiscence, provides good mechanical protection and shorten the time of anastomoses healing.

  2. Fibrin Sheath Angioplasty: A Technique to Prevent Superior Vena Cava Stenosis Secondary to Dialysis Catheters

    Science.gov (United States)

    Hacker, Robert I.; Garcia, Lorena De Marco; Chawla, Ankur; Panetta, Thomas F.

    2012-01-01

    Fibrin sheaths are a heterogeneous matrix of cells and debris that form around catheters and are a known cause of central venous stenosis and catheter failure. A total of 50 cases of central venous catheter fibrin sheath angioplasty (FSA) after catheter removal or exchange are presented. A retrospective review of an outpatient office database identified 70 eligible patients over a 19-month period. After informed consent was obtained, the dialysis catheter exiting the skin was clamped, amputated, and a wire was inserted. The catheter was then removed and a 9-French sheath was inserted into the superior vena cava, a venogram was performed. If a fibrin sheath was present, angioplasty was performed using an 8 × 4 or 10 × 4 balloon along the entire length of the fibrin sheath. A completion venogram was performed to document obliteration of the sheath. During the study, 50 patients were diagnosed with a fibrin sheath, and 43 had no pre-existing central venous stenosis. After FSA, 39 of the 43 patient's (91%) central systems remained patent without the need for subsequent interventions; 3 patients (7%) developed subclavian stenoses requiring repeat angioplasty and stenting; 1 patent (2.3%) developed an occlusion requiring a reintervention. Seven patients with prior central stenosis required multiple angioplasties; five required stenting of their central lesions. Every patient had follow-up fistulograms to document long-term patency. We propose that FSA is a prudent and safe procedure that may help reduce the risk of central venous stenosis from fibrin sheaths due to central venous catheters. PMID:23997555

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

    Directory of Open Access Journals (Sweden)

    S Capossela

    2014-04-01

    Full Text Available 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.

  4. Chondrogenic potential of physically treated bovine cartilage matrix derived porous scaffolds on human dermal fibroblast cells.

    Science.gov (United States)

    Moradi, Ali; Ataollahi, Forough; Sayar, Katayoun; Pramanik, Sumit; Chong, Pan-Pan; Khalil, Alizan Abdul; Kamarul, Tunku; Pingguan-Murphy, Belinda

    2016-01-01

    Extracellular matrices have drawn attention in tissue engineering as potential biomaterials for scaffold fabrication because of their bioactive components. Noninvasive techniques of scaffold fabrication and cross-linking treatments are believed to maintain the integrity of bioactive molecules while providing proper architectural and mechanical properties. Cartilage matrix derived scaffolds are designed to support the maintenance of chondrocytes and provide proper signals for differentiation of chondroinducible cells. Chondroinductive potential of bovine articular cartilage matrix derived porous scaffolds on human dermal fibroblasts and the effect of scaffold shrinkage on chondrogenesis were investigated. An increase in sulfated glycosaminoglycans production along with upregulation of chondrogenic genes confirmed that physically treated cartilage matrix derived scaffolds have chondrogenic potential on human dermal fibroblasts. © 2015 Wiley Periodicals, Inc.

  5. Stretching single fibrin fibers hampers their lysis.

    Science.gov (United States)

    Li, Wei; Lucioni, Tomas; Li, Rongzhong; Bonin, Keith; Cho, Samuel S; Guthold, Martin

    2017-09-15

    Blood clots, whose main structural component is a mesh of microscopic fibrin fibers, experience mechanical strain from blood flow, clot retraction and interactions with platelets and other cells. We developed a transparent, striated and highly stretchable substrate made from fugitive glue (a styrenic block copolymer) to investigate how mechanical strain affects lysis of single, suspended fibrin fibers. In this suspended fiber assay, lysis manifested itself by fiber elongation, thickening (disassembly), fraying and collapse. Stretching single fibrin fibers significantly hampered their lysis. This effect was seen in uncrosslinked and crosslinked fibers. Crosslinking (without stretching) also hampered single fiber lysis. Our data suggest that strain is a novel mechanosensitive factor that regulates blood clot dissolution (fibrinolysis) at the single fiber level. At the molecular level of single fibrin molecules, strain may distort, or hinder access to, plasmin cleavage sites and thereby hamper lysis. Fibrin fibers are the major structural component of a blood clot. We developed a highly stretchable substrate made from fugitive glue and a suspended fibrin fiber lysis assay to investigate the effect of stretching on single fibrin fibers lysis. The key findings from our experiments are: 1) Fibers thicken and elongate upon lysis; 2) stretching strongly reduces lysis; 3) this effect is more pronounced for uncrosslinked fibers; and 4) stretching fibers has a similar effect on reducing lysis as crosslinking fibers. At the molecular level, strain may distort plasmin cleavage sites, or restrict access to those sites. Our results suggest that strain may be a novel mechanobiological factor that regulates fibrinolysis. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  6. Antimicrobial drugs encapsulated in fibrin nanoparticles for treating microbial infested wounds.

    Science.gov (United States)

    Alphonsa, B Maria; Sudheesh Kumar, P T; Praveen, G; Biswas, Raja; Chennazhi, K P; Jayakumar, R

    2014-05-01

    In vitro evaluation of antibacterial and antifungal drugs encapsulated fibrin nanoparticles to prove their potential prospect of using these nanocomponent for effective treatment of microbial infested wounds. Surfactant-free oil-in-water emulsification-diffusion method was adopted to encapsulate 1 mg/ml each of antimicrobial drugs (Ciprofloxacin and Fluconazole) in 4 ml of aqueous fibrinogen suspension and subsequent thrombin mediated cross linking to synthesize drug loaded fibrin nanoparticles. Ciprofloxacin loaded fibrin nanoparticles (CFNPs) showed size range of 253 ± 6 nm whereas that of Fluconazole loaded fibrin nanoparticles (FFNPs) was 260 ± 10 nm. Physico chemical characterizations revealed the firm integration of antimicrobial drugs within fibrin nanoparticles. Drug release studies performed at physiological pH 7.4 showed a release of 16% ciprofloxacin and 8% of fluconazole while as the release of ciprofloxacin at alkaline pH 8.5, was 48% and that of fluconazole was 37%. The antimicrobial activity evaluations of both drug loaded systems independently showed good antibacterial activity against Escherichia coli (E.coli), Staphylococcus aureus (S. aureus) and antifungal activity against Candida albicans (C. albicans). The in vitro toxicity of the prepared drug loaded nanoparticles were further analyzed using Human dermal fibroblast cells (HDF) and showed adequate cell viability. The efficacies of both CFNPs and FFNPs for sustained delivery of encapsulated anti microbial drugs were evaluated in vitro suggesting its potential use for treating microbial infested wounds (diabetic foot ulcer).

  7. Human 2'-phosphodiesterase localizes to the mitochondrial matrix with a putative function in mitochondrial RNA turnover

    DEFF Research Database (Denmark)

    Poulsen, Jesper Buchhave; Andersen, Kasper Røjkjær; Kjær, Karina Hansen

    2011-01-01

    . Interestingly, 2′-PDE shares both functionally and structurally characteristics with the CCR4-type exonuclease–endonuclease–phosphatase family of deadenylases. Here we show that 2′-PDE locates to the mitochondrial matrix of human cells, and comprise an active 3′–5′ exoribonuclease exhibiting a preference...

  8. Alginate nanobeads interspersed fibrin network as in situ forming hydrogel for soft tissue engineering

    Directory of Open Access Journals (Sweden)

    S. Deepthi

    2018-06-01

    Full Text Available Hydrogels are a class of materials that has the property of injectability and in situ gel formation. This property of hydrogels is manipulated in this study to develop a biomimetic bioresorbable injectable system of alginate nanobeads interspersed in fibrin network. Alginate nanobeads developed by calcium cross-linking yielded a size of 200–500 nm. The alginate nanobeads fibrin hydrogel was formed using dual syringe apparatus. Characterization of the in situ injectable hydrogel was done by SEM, FTIR and Rheometer. The developed hydrogel showed mechanical strength of 19 kPa which provides the suitable compliance for soft tissue engineering. Cytocompatibility studies using human umbilical cord blood derived mesenchymal stem cells showed good attachment, proliferation and infiltration within the hydrogel similar to fibrin gel. The developed in situ forming hydrogel could be a suitable delivery carrier of stem cells for soft tissue regeneration.

  9. Alginate nanobeads interspersed fibrin network as in situ forming hydrogel for soft tissue engineering.

    Science.gov (United States)

    Deepthi, S; Jayakumar, R

    2018-06-01

    Hydrogels are a class of materials that has the property of injectability and in situ gel formation. This property of hydrogels is manipulated in this study to develop a biomimetic bioresorbable injectable system of alginate nanobeads interspersed in fibrin network. Alginate nanobeads developed by calcium cross-linking yielded a size of 200-500 nm. The alginate nanobeads fibrin hydrogel was formed using dual syringe apparatus. Characterization of the in situ injectable hydrogel was done by SEM, FTIR and Rheometer. The developed hydrogel showed mechanical strength of 19 kPa which provides the suitable compliance for soft tissue engineering. Cytocompatibility studies using human umbilical cord blood derived mesenchymal stem cells showed good attachment, proliferation and infiltration within the hydrogel similar to fibrin gel. The developed in situ forming hydrogel could be a suitable delivery carrier of stem cells for soft tissue regeneration.

  10. Postsynaptic density protein 95 in the striosome and matrix compartments of the human neostriatum.

    Directory of Open Access Journals (Sweden)

    Ryoma eMorigaki

    2015-11-01

    Full Text Available The human neostriatum consists of two functional subdivisions referred to as the striosome (patch and matrix compartments. The striosome-matrix dopamine systems play a central role in cortico-thalamo-basal ganglia circuits, and their involvement is thought to underlie the genesis of multiple movement and behavioral disorders, and of drug addiction. Human neuropathology also has shown that striosomes and matrix have differential vulnerability patterns in several striatal neurodegenerative diseases. Postsynaptic density protein 95 (PSD-95, also known as DLG4, is a major scaffolding protein in the postsynaptic densities of dendritic spines. PSD-95 is now known to negatively regulate not only N-methyl-D-aspartate glutamate signaling, but also dopamine D1 signals at sites of postsynaptic transmission. Accordingly, a neuroprotective role for PSD-95 against dopamine D1 receptor (D1R-mediated neurotoxicity in striatal neurodegeneration also has been suggested. Here, we used a highly sensitive immunohistochemistry technique to show that in the human neostriatum, PSD-95 is differentially concentrated in the striosome and matrix compartments, with a higher density of PSD-95 labeling in the matrix compartment than in the striosomes. This compartment-specific distribution of PSD-95 was strikingly complementary to that of D1R. In addition to the possible involvement of PSD-95-mediated synaptic function in compartment-specific dopamine signals, we suggest that the striosomes might be more susceptible to D1R-mediated neurotoxicity than the matrix compartment. This notion may provide new insight into the compartment-specific vulnerability of MSNs in striatal neurodegeneration.

  11. Nonexpansive immediate breast reconstruction using human acellular tissue matrix graft (AlloDerm).

    Science.gov (United States)

    Salzberg, C Andrew

    2006-07-01

    Immediate breast reconstruction has become a standard of care following mastectomy for cancer, largely due to improved esthetic and psychologic outcomes achieved with this technique. However, the current historical standards--transverse rectus abdominis myocutaneous flap reconstruction and expander--implant surgery-still have limitations as regards patient morbidity, short-term body-image improvements, and even cost. To address these shortcomings, we employ a novel concept of human tissue replacement to enhance breast shape and provide total coverage, enabling immediate mound reconstruction without the need for breast expansion prior to permanent implant placement. AlloDerm (human acellular tissue matrix) is a human-derived graft tissue with extensive experience in various settings of skin and soft tissue replacement surgery. This report describes the success using acellular tissue matrix to provide total coverage over the prosthesis in immediate reconstruction, with limited muscle dissection. In this population, 49 patients (76 breasts) successfully underwent the acellular tissue matrix-based immediate reconstruction, resulting in durable breast reconstruction with good symmetry. These findings may predict that acellular tissue matrix-supplemented immediate breast reconstruction will become a new technique for the immediate reconstruction of the postmastectomy breast.

  12. Chondrocyte secreted CRTAC1: a glycosylated extracellular matrix molecule of human articular cartilage.

    Science.gov (United States)

    Steck, Eric; Bräun, Jessica; Pelttari, Karoliina; Kadel, Stephanie; Kalbacher, Hubert; Richter, Wiltrud

    2007-01-01

    Cartilage acidic protein 1 (CRTAC1), a novel human marker which allowed discrimination of human chondrocytes from osteoblasts and mesenchymal stem cells in culture was so far studied only on the RNA-level. We here describe its genomic organisation and detect a new brain expressed (CRTAC1-B) isoform resulting from alternate last exon usage which is highly conserved in vertebrates. In humans, we identify an exon sharing process with the neighbouring tail-to-tail orientated gene leading to CRTAC1-A. This isoform is produced by cultured human chondrocytes, localized in the extracellular matrix of articular cartilage and its secretion can be stimulated by BMP4. Of five putative O-glycosylation motifs in the last exon of CRTAC1-A, the most C-terminal one is modified according to exposure of serial C-terminal deletion mutants to the O-glycosylation inhibitor Benzyl-alpha-GalNAc. Both isoforms contain four FG-GAP repeat domains and an RGD integrin binding motif, suggesting cell-cell or cell-matrix interaction potential. In summary, CRTAC1 acquired an alternate last exon from the tail-to-tail oriented neighbouring gene in humans resulting in the glycosylated isoform CRTAC1-A which represents a new extracellular matrix molecule of articular cartilage.

  13. A Human Nuclear Shuttling Protein That Interacts with Human Immunodeficiency Virus Type 1 Matrix Is Packaged into Virions

    Science.gov (United States)

    Gupta, Kalpana; Ott, David; Hope, Thomas J.; Siliciano, Robert F.; Boeke, Jef D.

    2000-01-01

    Active nuclear import of the human immunodeficiency virus type 1 (HIV-1) preintegration complex (PIC) is essential for the productive infection of nondividing cells. Nuclear import of the PIC is mediated by the HIV-1 matrix protein, which also plays several critical roles during viral entry and possibly during virion production facilitating the export of Pr55Gag and genomic RNA. Using a yeast two-hybrid screen, we identified a novel human virion-associated matrix-interacting protein (VAN) that is highly conserved in vertebrates and expressed in most human tissues. Its expression is upregulated upon activation of CD4+ T cells. VAN is efficiently incorporated into HIV-1 virions and, like matrix, shuttles between the nucleus and cytoplasm. Furthermore, overexpression of VAN significantly inhibits HIV-1 replication in tissue culture. We propose that VAN regulates matrix nuclear localization and, by extension, both nuclear import of the PIC and export of Pr55Gag and viral genomic RNA during virion production. Our data suggest that this regulatory mechanism reflects a more global process for regulation of nucleocytoplasmic transport. PMID:11090181

  14. Matrix Stiffness Corresponding to Strictured Bowel Induces a Fibrogenic Response in Human Colonic Fibroblasts

    Science.gov (United States)

    Johnson, Laura A.; Rodansky, Eva S.; Sauder, Kay L.; Horowitz, Jeffrey C.; Mih, Justin D.; Tschumperlin, Daniel J.; Higgins, Peter D.

    2013-01-01

    Background Crohn’s disease is characterized by repeated cycles of inflammation and mucosal healing which ultimately progress to intestinal fibrosis. This inexorable progression towards fibrosis suggests that fibrosis becomes inflammation-independent and auto-propagative. We hypothesized that matrix stiffness regulates this auto-propagation of intestinal fibrosis. Methods The stiffness of fresh ex vivo samples from normal human small intestine, Crohn’s disease strictures, and the unaffected margin were measured with a microelastometer. Normal human colonic fibroblasts were cultured on physiologically normal or pathologically stiff matrices corresponding to the physiological stiffness of normal or fibrotic bowel. Cellular response was assayed for changes in cell morphology, α-smooth muscle actin (αSMA) staining, and gene expression. Results Microelastometer measurements revealed a significant increase in colonic tissue stiffness between normal human colon and Crohn’s strictures as well as between the stricture and adjacent tissue margin. In Ccd-18co cells grown on stiff matrices corresponding to Crohn’s strictures, cellular proliferation increased. Pathologic stiffness induced a marked change in cell morphology and increased αSMA protein expression. Growth on a stiff matrix induced fibrogenic gene expression, decreased matrix metalloproteinase and pro-inflammatory gene expression, and was associated with nuclear localization of the transcriptional cofactor MRTF-A. Conclusions Matrix stiffness, representative of the pathological stiffness of Crohn’s strictures, activates human colonic fibroblasts to a fibrogenic phenotype. Matrix stiffness affects multiple pathways suggesting the mechanical properties of the cellular environment are critical to fibroblast function and may contribute to autopropagation of intestinal fibrosis in the absence of inflammation, thereby contributing to the intractable intestinal fibrosis characteristic of Crohn’s disease. PMID

  15. In vitro human periodontal ligament-like tissue formation with porous poly-L-lactide matrix

    International Nuclear Information System (INIS)

    Liao, Wen; Okada, Masahiro; Sakamoto, Fumito; Okita, Naoya; Inami, Kaoru; Nishiura, Aki; Hashimoto, Yoshiya; Matsumoto, Naoyuki

    2013-01-01

    This study aimed to establish an in vitro human periodontal ligament-like tissue (HPdLLT) by three-dimensional culturing of human periodontal ligament fibroblasts (HPdLFs) in a porous poly-L-lactide (PLLA) matrix modified hydrophilically with ammonia solution. After ammonia modification, the surface roughness and culture-medium-soaking-up ability of the PLLA matrix increased, whereas the contact angle of water drops decreased. The thickness, porosity, and pore size of the PLLA matrix were 400 ± 50 μm, 83.3%, and 75–150 μm, respectively. HPdLFs (1 × 10 5 cells) were seeded on the modified PLLA matrix and centrifuged to facilitate seeding into its interior and cultured for 14 days. Scanning electron microscope (SEM) observation, proliferation assay, picrosirius-red staining, and real-time polymerase chain reaction (RT-PCR) for type-1 collagen (COL1), periodontal ligament associated protein-1 (PLAP-1), fibroblast growth factor-2 (FGF-2), and alkaline phosphatase (ALP) mRNA were conducted on days 1, 3, 7, and 14. HPdLFs were observed entirely from the surface to the rear side of the matrix. Cell proliferation analysis, SEM observation, and picrosirius-red staining showed both progressive growth of 3D-cultured HPdLFs and extracellular matrix maturation by the secretion of COL1 and type 3 collagen (COL3) from days 1 to 14. Expressions of COL1, PLAP-1, and FGF-2 mRNA suggested the formation of cellular components and supplementation of extracellular components. Expressions of ALP, COL1, and PLAP-1 mRNA suggested the osteogenic potential of the HPdLLT. The results indicated in vitro HPdLLT formation, and it could be used in future periodontal ligament tissue engineering to achieve optimal periodontal regeneration. - Highlights: • First report on ammonia treated PLLA matrix for in vitro human periodontal ligament-like tissue generation. • Good combination of matrix thickness, pore size, and porosity. • Biodegradable PLLA is also possible to be used in vivo

  16. In vitro human periodontal ligament-like tissue formation with porous poly-L-lactide matrix

    Energy Technology Data Exchange (ETDEWEB)

    Liao, Wen [Graduate School of Dentistry, Department of Orthodontics, Osaka Dental University, 8-1 Kuzuha-hanazono-cho, Hirakata-shi, Osaka-fu 573-1121 (Japan); Okada, Masahiro [Department of Biomaterials, Osaka Dental University, 8-1 Kuzuha-hanazono-cho, Hirakata-shi, Osaka-fu 573-1121 (Japan); Sakamoto, Fumito; Okita, Naoya [Graduate School of Dentistry, Department of Orthodontics, Osaka Dental University, 8-1 Kuzuha-hanazono-cho, Hirakata-shi, Osaka-fu 573-1121 (Japan); Inami, Kaoru; Nishiura, Aki [Department of Orthodontics, Osaka Dental University, 8-1 Kuzuha-hanazono-cho, Hirakata-shi, Osaka-fu 573-1121 (Japan); Hashimoto, Yoshiya, E-mail: yoshiya@cc.osaka-dent.ac.jp [Department of Biomaterials, Osaka Dental University, 8-1 Kuzuha-hanazono-cho, Hirakata-shi, Osaka-fu 573-1121 (Japan); Matsumoto, Naoyuki [Department of Orthodontics, Osaka Dental University, 8-1 Kuzuha-hanazono-cho, Hirakata-shi, Osaka-fu 573-1121 (Japan)

    2013-08-01

    This study aimed to establish an in vitro human periodontal ligament-like tissue (HPdLLT) by three-dimensional culturing of human periodontal ligament fibroblasts (HPdLFs) in a porous poly-L-lactide (PLLA) matrix modified hydrophilically with ammonia solution. After ammonia modification, the surface roughness and culture-medium-soaking-up ability of the PLLA matrix increased, whereas the contact angle of water drops decreased. The thickness, porosity, and pore size of the PLLA matrix were 400 ± 50 μm, 83.3%, and 75–150 μm, respectively. HPdLFs (1 × 10{sup 5} cells) were seeded on the modified PLLA matrix and centrifuged to facilitate seeding into its interior and cultured for 14 days. Scanning electron microscope (SEM) observation, proliferation assay, picrosirius-red staining, and real-time polymerase chain reaction (RT-PCR) for type-1 collagen (COL1), periodontal ligament associated protein-1 (PLAP-1), fibroblast growth factor-2 (FGF-2), and alkaline phosphatase (ALP) mRNA were conducted on days 1, 3, 7, and 14. HPdLFs were observed entirely from the surface to the rear side of the matrix. Cell proliferation analysis, SEM observation, and picrosirius-red staining showed both progressive growth of 3D-cultured HPdLFs and extracellular matrix maturation by the secretion of COL1 and type 3 collagen (COL3) from days 1 to 14. Expressions of COL1, PLAP-1, and FGF-2 mRNA suggested the formation of cellular components and supplementation of extracellular components. Expressions of ALP, COL1, and PLAP-1 mRNA suggested the osteogenic potential of the HPdLLT. The results indicated in vitro HPdLLT formation, and it could be used in future periodontal ligament tissue engineering to achieve optimal periodontal regeneration. - Highlights: • First report on ammonia treated PLLA matrix for in vitro human periodontal ligament-like tissue generation. • Good combination of matrix thickness, pore size, and porosity. • Biodegradable PLLA is also possible to be used in vivo.

  17. Subfractions of enamel matrix derivative differentially influence cytokine secretion from human oral fibroblasts

    Directory of Open Access Journals (Sweden)

    Oscar Villa

    2015-03-01

    Full Text Available Enamel matrix derivative is used to promote periodontal regeneration during the corrective phase of the treatment of periodontal defects. Our main goal was to analyze the bioactivity of different molecular weight fractions of enamel matrix derivative. Enamel matrix derivative, a complex mixture of proteins, was separated into 13 fractions using size-exclusion chromatography and characterized by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and liquid chromatography–electrospray ionization–tandem mass spectrometry. Human periodontal ligament fibroblasts were treated with either enamel matrix derivative or the different fractions. Proliferation and cytokine secretion to the cell culture medium were measured and compared to untreated cells. The liquid chromatography–electrospray ionization–tandem mass spectrometry analyses revealed that the most abundant peptides were amelogenin and leucine-rich amelogenin peptide related. The fractions containing proteins above 20 kDa induced an increase in vascular endothelial growth factor and interleukin-6 secretion, whereas lower molecular weight fractions enhanced proliferation and secretion of interleukin-8 and monocyte chemoattractant protein-1 and reduced interleukin-4 release. The various molecular components in the enamel matrix derivative formulation might contribute to reported effects on tissue regeneration through their influence on vascularization, the immune response, and chemotaxis.

  18. Growth of hydroxyapatite on physiologically clotted fibrin capped gold nanoparticles

    International Nuclear Information System (INIS)

    Sastry, T P; Sundaraseelan, J; Swarnalatha, K; Sobhana, S S Liji; Makheswari, M Uma; Sekar, S; Mandal, A B

    2008-01-01

    The growth of hydroxyapatite (HAp) on physiologically clotted fibrin (PCF)-gold nanoparticles is presented for the first time by employing a wet precipitation method. Fourier transform infrared (FTIR) spectroscopy confirmed the characteristic functionalities of PCF and HAp in the PCF-Au-HAp nanocomposite. Scanning electron microscopy (SEM) images have shown cuboidal nanostructures having a size in the range of 70-300 nm of HAp, whereas 2-50 nm sized particles were visualized in high-resolution transmission electron microscopy (TEM). Energy-dispersive x-ray (EDX) and x-ray diffraction (XRD) studies have confirmed the presence of HAp. These results show that gold nanoparticles with PCF acted as a matrix for the growth of HAp, and that PCF-Au-HAp nanocomposite is expected to have better osteoinductive properties

  19. Decellularized Matrix from Tumorigenic Human Mesenchymal Stem Cells Promotes Neovascularization with Galectin-1 Dependent Endothelial Interaction

    Science.gov (United States)

    Burns, Jorge S.; Kristiansen, Malthe; Kristensen, Lars P.; Larsen, Kenneth H.; Nielsen, Maria O.; Christiansen, Helle; Nehlin, Jan; Andersen, Jens S.; Kassem, Moustapha

    2011-01-01

    Background Acquisition of a blood supply is fundamental for extensive tumor growth. We recently described vascular heterogeneity in tumours derived from cell clones of a human mesenchymal stem cell (hMSC) strain (hMSC-TERT20) immortalized by retroviral vector mediated human telomerase (hTERT) gene expression. Histological analysis showed that cells of the most vascularized tumorigenic clone, -BD11 had a pericyte-like alpha smooth muscle actin (ASMA+) and CD146+ positive phenotype. Upon serum withdrawal in culture, -BD11 cells formed cord-like structures mimicking capillary morphogenesis. In contrast, cells of the poorly tumorigenic clone, -BC8 did not stain for ASMA, tumours were less vascularized and serum withdrawal in culture led to cell death. By exploring the heterogeneity in hMSC-TERT20 clones we aimed to understand molecular mechanisms by which mesenchymal stem cells may promote neovascularization. Methodology/Principal Findings Quantitative qRT-PCR analysis revealed similar mRNA levels for genes encoding the angiogenic cytokines VEGF and Angiopoietin-1 in both clones. However, clone-BD11 produced a denser extracellular matrix that supported stable ex vivo capillary morphogenesis of human endothelial cells and promoted in vivo neovascularization. Proteomic characterization of the -BD11 decellularized matrix identified 50 extracellular angiogenic proteins, including galectin-1. siRNA knock down of galectin-1 expression abrogated the ex vivo interaction between decellularized -BD11 matrix and endothelial cells. More stable shRNA knock down of galectin-1 expression did not prevent -BD11 tumorigenesis, but greatly reduced endothelial migration into -BD11 cell xenografts. Conclusions Decellularized hMSC matrix had significant angiogenic potential with at least 50 angiogenic cell surface and extracellular proteins, implicated in attracting endothelial cells, their adhesion and activation to form tubular structures. hMSC -BD11 surface galectin-1 expression was

  20. Neutrophil-Derived MMP-8 Drives AMPK-Dependent Matrix Destruction in Human Pulmonary Tuberculosis

    Science.gov (United States)

    Ong, Catherine W. M.; Elkington, Paul T.; Brilha, Sara; Ugarte-Gil, Cesar; Tome-Esteban, Maite T.; Tezera, Liku B.; Pabisiak, Przemyslaw J.; Moores, Rachel C.; Sathyamoorthy, Tarangini; Patel, Vimal; Gilman, Robert H.; Porter, Joanna C.; Friedland, Jon S.

    2015-01-01

    Pulmonary cavities, the hallmark of tuberculosis (TB), are characterized by high mycobacterial load and perpetuate the spread of M. tuberculosis. The mechanism of matrix destruction resulting in cavitation is not well defined. Neutrophils are emerging as key mediators of TB immunopathology and their influx are associated with poor outcomes. We investigated neutrophil-dependent mechanisms involved in TB-associated matrix destruction using a cellular model, a cohort of 108 patients, and in separate patient lung biopsies. Neutrophil-derived NF-kB-dependent matrix metalloproteinase-8 (MMP-8) secretion was up-regulated in TB and caused matrix destruction both in vitro and in respiratory samples of TB patients. Collagen destruction induced by TB infection was abolished by doxycycline, a licensed MMP inhibitor. Neutrophil extracellular traps (NETs) contain MMP-8 and are increased in samples from TB patients. Neutrophils lined the circumference of human pulmonary TB cavities and sputum MMP-8 concentrations reflected TB radiological and clinical disease severity. AMPK, a central regulator of catabolism, drove neutrophil MMP-8 secretion and neutrophils from AMPK-deficient patients secrete lower MMP-8 concentrations. AMPK-expressing neutrophils are present in human TB lung biopsies with phospho-AMPK detected in nuclei. These data demonstrate that neutrophil-derived MMP-8 has a key role in the immunopathology of TB and is a potential target for host-directed therapy in this infectious disease. PMID:25996154

  1. Neutrophil-Derived MMP-8 Drives AMPK-Dependent Matrix Destruction in Human Pulmonary Tuberculosis.

    Science.gov (United States)

    Ong, Catherine W M; Elkington, Paul T; Brilha, Sara; Ugarte-Gil, Cesar; Tome-Esteban, Maite T; Tezera, Liku B; Pabisiak, Przemyslaw J; Moores, Rachel C; Sathyamoorthy, Tarangini; Patel, Vimal; Gilman, Robert H; Porter, Joanna C; Friedland, Jon S

    2015-05-01

    Pulmonary cavities, the hallmark of tuberculosis (TB), are characterized by high mycobacterial load and perpetuate the spread of M. tuberculosis. The mechanism of matrix destruction resulting in cavitation is not well defined. Neutrophils are emerging as key mediators of TB immunopathology and their influx are associated with poor outcomes. We investigated neutrophil-dependent mechanisms involved in TB-associated matrix destruction using a cellular model, a cohort of 108 patients, and in separate patient lung biopsies. Neutrophil-derived NF-kB-dependent matrix metalloproteinase-8 (MMP-8) secretion was up-regulated in TB and caused matrix destruction both in vitro and in respiratory samples of TB patients. Collagen destruction induced by TB infection was abolished by doxycycline, a licensed MMP inhibitor. Neutrophil extracellular traps (NETs) contain MMP-8 and are increased in samples from TB patients. Neutrophils lined the circumference of human pulmonary TB cavities and sputum MMP-8 concentrations reflected TB radiological and clinical disease severity. AMPK, a central regulator of catabolism, drove neutrophil MMP-8 secretion and neutrophils from AMPK-deficient patients secrete lower MMP-8 concentrations. AMPK-expressing neutrophils are present in human TB lung biopsies with phospho-AMPK detected in nuclei. These data demonstrate that neutrophil-derived MMP-8 has a key role in the immunopathology of TB and is a potential target for host-directed therapy in this infectious disease.

  2. Complete horizontal skin cell resurfacing and delayed vertical cell infiltration into porcine reconstructive tissue matrix compared to bovine collagen matrix and human dermis.

    Science.gov (United States)

    Mirastschijski, Ursula; Kerzel, Corinna; Schnabel, Reinhild; Strauss, Sarah; Breuing, Karl-Heinz

    2013-10-01

    Xenogenous dermal matrices are used for hernia repair and breast reconstruction. Full-thickness skin replacement is needed after burn or degloving injuries with exposure of tendons or bones. The authors used a human skin organ culture model to study whether porcine reconstructive tissue matrix (Strattice) is effective as a dermal tissue replacement. Skin cells or split-thickness skin grafts were seeded onto human deepidermized dermis, Strattice, and Matriderm. Cellular resurfacing and matrix infiltration were monitored by live fluorescence imaging, histology, and electron microscopy. Proliferation, apoptosis, cell differentiation, and adhesion were analyzed by immunohistochemistry. Epithelial resurfacing and vertical proliferation were reduced and delayed with both bioartificial matrices compared with deepidermized dermis; however, no differences in apoptosis, cell differentiation, or basement membrane formation were found. Vertical penetration was greatest on Matriderm, whereas no matrix infiltration was found on Strattice in the first 12 days. Uncompromised horizontal resurfacing was greatest with Strattice but was absent with Matriderm. Strattice showed no stimulatory effect on cellular inflammation. Matrix texture and surface properties governed cellular performance on tissues. Although dense dermal compaction delayed vertical cellular ingrowth for Strattice, it allowed uncompromised horizontal resurfacing. Dense dermal compaction may slow matrix decomposition and result in prolonged biomechanical stability of the graft. Reconstructive surgeons should choose the adequate matrix substitute depending on biomechanical requirements at the recipient site. Strattice may be suitable as a dermal replacement at recipient sites with high mechanical load requirements.

  3. Cell-seeded polyurethane-fibrin structures – A possible system for intervertebral disc regeneration

    Directory of Open Access Journals (Sweden)

    C Mauth

    2009-10-01

    Full Text Available Nowadays, intervertebral disc (IVD degeneration is one of the principal causes of low back pain involving high expense within the health care system. The long-term goal is the development of a medical treatment modality focused on a more biological regeneration of the inner nucleus pulposus (NP. Hence, interest in the endoscopic implantation of an injectable material took center stage in the recent past. We report on the development of a novel polyurethane (PU scaffold as a mechanically stable carrier system for the reimplantation of expanded autologous IVD-derived cells (disc cells to stimulate regenerative processes and restore the chondrocyte-like tissue within the NP. Primary human disc cells were seeded into newly developed PU spheroids which were subsequently encapsulated in fibrin hydrogel. The study aims to analyze adhesion properties, proliferation capacity and phenotypic characterization of these cells. Polymerase chain reaction was carried out to detect the expression of genes specifically expressed by native IVD cells. Biochemical analyses showed an increased DNA content, and a progressive enhancement of total collagen and glycosaminoglycans (GAG was observed during cell culture. The results suggest the synthesis of an appropriate extracellular matrix as well as a stable mRNA expression of chondrogenic and/or NP specific markers. In conclusion, the data presented indicate an alternative medical approach to current treatment options of degenerated IVD tissue.

  4. Culture on fibrin matrices maintains the colony-forming capacity and osteoblastic differentiation of mesenchymal stem cells

    International Nuclear Information System (INIS)

    Colley, Helen; McArthur, Sally L; Stolzing, Alexandra; Scutt, Andy

    2012-01-01

    Mesenchymal stem cells (MSC) are multipotent cells capable of differentiating into a number of mesenchymal tissues including bone, cartilage, and tendon. Low numbers in vivo means exponential growth is needed in culture to enable therapeutic applications. MSC can expand rapidly in culture but usually lose their extensive capacity for differentiation that makes them therapeutically attractive. To try and maintain their capacity for differentiation and expansion in vitro, we cultured MSC on fibrin gels of different concentrations to create more physiological growth conditions for the cells. The cells were then re-plated onto tissue culture plastic and analysed. The cells that had been pre-cultured for seven days on fibrin, proliferated and maintained their differential potential to the osteogenic lineage better than tissue culture plastic expanded MSC. A concentration relationship between colony number and fibrin concentration was seen with decreasing numbers as fibrin concentration increased. These data support the concept that substrate signals significantly influence MSC growth and differentiation and that growth on a fibrin matrix could be used to maintain a stem cell phenotype during MSC expansion. (paper)

  5. Three-dimensional architecture and cell composition of a Choukroun's platelet-rich fibrin clot and membrane.

    Science.gov (United States)

    Dohan Ehrenfest, David M; Del Corso, Marco; Diss, Antoine; Mouhyi, Jaafar; Charrier, Jean-Baptiste

    2010-04-01

    Platelet-rich fibrin (PRF; Choukroun's technique) is a second-generation platelet concentrate for surgical use. This easy protocol allows the production of leukocyte and platelet-rich fibrin clots and membranes starting from 10-ml blood samples. The purposes of this study were to determine the cell composition and three-dimensional organization of this autologous biomaterial and to evaluate the influence of different collection tubes (dry glass or glass-coated plastic tubes) and compression procedures (forcible or soft) on the final PRF-membrane architecture. After centrifugation, blood analyses were performed on the residual waste plasmatic layers after collecting PRF clots. The PRF clots and membranes were processed for examination by light microscopy and scanning electron microscopy. Approximately 97% of the platelets and >50% of the leukocytes were concentrated in the PRF clot and showed a specific three-dimensional distribution, depending on the centrifugation forces. Platelets and fibrin formed large clusters of coagulation in the first millimeters of the membrane beyond the red blood cell base. The fibrin network was very mature and dense. Moreover, there was no significant difference in the PRF architecture between groups using the different tested collection tubes and compression techniques, even if these two parameters could have influenced the growth factor content and biologic matrix properties. The PRF protocol concentrated most platelets and leukocytes from a blood harvest into a single autologous fibrin biomaterial. This protocol offers reproducible results as long as the main production principles are respected.

  6. Platelet-rich fibrin: the benefits.

    Science.gov (United States)

    Kumar, Yuvika Raj; Mohanty, Sujata; Verma, Mahesh; Kaur, Raunaq Reet; Bhatia, Priyanka; Kumar, Varun Raj; Chaudhary, Zainab

    2016-01-01

    Current published data presents confusing results about the effects of platelet-rich fibrin on bone, and there is a need for studies that throw light on its effect. Our main objective therefore was to evaluate (by fractal analysis) osseous regeneration in extraction sockets with and without platelet-rich fibrin in a study with a substantial sample and a reliable technique to calibrate its effects on bone cells. We also assessed the soft tissue response. Thirty-four patients had their bilaterally impacted third molars (68 surgical sites) extracted in this split-mouth study, following which platelet-rich fibrin was placed in one of the sockets. Patients were followed up clinically and radiographically, and a pain score and fractal analysis were used to evaluate healing of soft tissue and bone, respectively. We conclude that platelet-rich fibrin improves healing of both soft and hard tissues. Although osseous healing did not differ significantly between the groups, healing of soft tissue as judged by the pain score was significantly better in the experimental group. Copyright © 2015 The British Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

  7. Differential Mueller matrix polarimetry technique for non-invasive measurement of glucose concentration on human fingertip.

    Science.gov (United States)

    Phan, Quoc-Hung; Lo, Yu-Lung

    2017-06-26

    A differential Mueller matrix polarimetry technique is proposed for obtaining non-invasive (NI) measurements of the glucose concentration on the human fingertip. The feasibility of the proposed method is demonstrated by detecting the optical rotation angle and depolarization index of tissue phantom samples containing de-ionized water (DI), glucose solutions with concentrations ranging from 0~500 mg/dL and 2% lipofundin. The results show that the extracted optical rotation angle increases linearly with an increasing glucose concentration, while the depolarization index decreases. The practical applicability of the proposed method is demonstrated by measuring the optical rotation angle and depolarization index properties of the human fingertips of healthy volunteers.

  8. Human endothelium on vascular prostheses modified by extracellular matrix proteins in a flow experiment

    Czech Academy of Sciences Publication Activity Database

    Chlupáč, Jaroslav; Filová, Elena; Riedel, Tomáš; Brynda, Eduard; Remy-Zolghadri, M.; Bareille, R.; Fernandez, P.; Daculsi, R.; Bordenave, L.; Bačáková, Lucie

    2006-01-01

    Roč. 9, č. 58-60 (2006), s. 10-13 ISSN 1429-7248 R&D Projects: GA AV ČR(CZ) IAA5011301; GA AV ČR(CZ) IAA4050202; GA AV ČR(CZ) IAA400500507; GA AV ČR(CZ) 1QS500110564 Institutional research plan: CEZ:AV0Z50110509; CEZ:AV0Z40500505 Keywords : bioartificial vascular prostheses * laminin * fibrin Subject RIV: EI - Biotechnology ; Bionics

  9. Fibrin and poly(lactic-co-glycolic acid) hybrid scaffold promotes early chondrogenesis of articular chondrocytes: an in vitro study.

    Science.gov (United States)

    Sha'ban, Munirah; Kim, Soon Hee; Idrus, Ruszymah Bh; Khang, Gilson

    2008-04-25

    Synthetic- and naturally derived- biodegradable polymers have been widely used to construct scaffolds for cartilage tissue engineering. Poly(lactic-co-glycolic acid) (PLGA) are bioresorbable and biocompatible, rendering them as a promising tool for clinical application. To minimize cells lost during the seeding procedure, we used the natural polymer fibrin to immobilize cells and to provide homogenous cells distribution in PLGA scaffolds. We evaluated in vitro chondrogenesis of rabbit articular chondrocytes in PLGA scaffolds using fibrin as cell transplantation matrix. PLGA scaffolds were soaked in chondrocytes-fibrin suspension (1 x 10(6) cells/scaffold) and polymerized by dropping thrombin-calcium chloride (CaCl2) solution. PLGA-seeded chondrocytes was used as control. All constructs were cultured for a maximum of 21 days. Cell proliferation activity was measured at 1, 3, 7, 14 and 21 days in vitro using 3-(4,5-dimethylthiazole-2-yl)-2-, 5-diphenyltetrazolium-bromide (MTT) assay. Morphological observation, histology, immunohistochemistry (IHC), gene expression and sulphated-glycosaminoglycan (sGAG) analyses were performed at each time point of 1, 2 and 3 weeks to elucidate in vitro cartilage development and deposition of cartilage-specific extracellular matrix (ECM). Cell proliferation activity was gradually increased from day-1 until day-14 and declined by day-21. A significant cartilaginous tissue formation was detected as early as 2-week in fibrin/PLGA hybrid construct as confirmed by the presence of cartilage-isolated cells and lacunae embedded within basophilic ECM. Cartilage formation was remarkably evidenced after 3 weeks. Presence of cartilage-specific proteoglycan and glycosaminoglycan (GAG) in fibrin/PLGA hybrid constructs were confirmed by positive Safranin O and Alcian Blue staining. Collagen type II exhibited intense immunopositivity at the pericellular matrix. Chondrogenic properties were further demonstrated by the expression of genes encoded for

  10. Fibrin and poly(lactic-co-glycolic acid hybrid scaffold promotes early chondrogenesis of articular chondrocytes: an in vitro study

    Directory of Open Access Journals (Sweden)

    Idrus Ruszymah BH

    2008-04-01

    Full Text Available Abstract Background Synthetic- and naturally derived- biodegradable polymers have been widely used to construct scaffolds for cartilage tissue engineering. Poly(lactic-co-glycolic acid (PLGA are bioresorbable and biocompatible, rendering them as a promising tool for clinical application. To minimize cells lost during the seeding procedure, we used the natural polymer fibrin to immobilize cells and to provide homogenous cells distribution in PLGA scaffolds. We evaluated in vitro chondrogenesis of rabbit articular chondrocytes in PLGA scaffolds using fibrin as cell transplantation matrix. Methods PLGA scaffolds were soaked in chondrocytes-fibrin suspension (1 × 106cells/scaffold and polymerized by dropping thrombin-calcium chloride (CaCl2 solution. PLGA-seeded chondrocytes was used as control. All constructs were cultured for a maximum of 21 days. Cell proliferation activity was measured at 1, 3, 7, 14 and 21 days in vitro using 3-(4,5-dimethylthiazole-2-yl-2-, 5-diphenyltetrazolium-bromide (MTT assay. Morphological observation, histology, immunohistochemistry (IHC, gene expression and sulphated-glycosaminoglycan (sGAG analyses were performed at each time point of 1, 2 and 3 weeks to elucidate in vitro cartilage development and deposition of cartilage-specific extracellular matrix (ECM. Results Cell proliferation activity was gradually increased from day-1 until day-14 and declined by day-21. A significant cartilaginous tissue formation was detected as early as 2-week in fibrin/PLGA hybrid construct as confirmed by the presence of cartilage-isolated cells and lacunae embedded within basophilic ECM. Cartilage formation was remarkably evidenced after 3 weeks. Presence of cartilage-specific proteoglycan and glycosaminoglycan (GAG in fibrin/PLGA hybrid constructs were confirmed by positive Safranin O and Alcian Blue staining. Collagen type II exhibited intense immunopositivity at the pericellular matrix. Chondrogenic properties were further

  11. Effects of matrix elasticity and cell density on human mesenchymal stem cells differentiation.

    Science.gov (United States)

    Xue, Ruyue; Li, Julie Yi-Shuan; Yeh, Yiting; Yang, Li; Chien, Shu

    2013-09-01

    Human mesenchymal stem cells (hMSCs) can differentiate into various cell types, including osteogenic and chondrogenic cells. The matrix elasticity and cell seeding density are important factors in hMSCs differentiation. We cultured hMSCs at different seeding densities on polyacrylamide hydrogels with different stiffness corresponding to Young's moduli of 1.6 ± 0.3 and 40 ± 3.6 kPa. The promotion of osteogenic marker expression by hard gel is overridden by a high seeding density. Cell seeding density, however, did not influence the chondrogenic marker expressions induced by soft gel. These findings suggest that interplays between cell-matrix and cell-cell interactions contribute to hMSCs differentiation. The promotion of osteogenic differentiation on hard matrix was shown to be mediated through the Ras pathway. Inhibition of Ras (RasN17) significantly decreased ERK, Smad1/5/8 and AKT activation, and osteogenic markers expression. However, constitutively active Ras (RasV12) had little effect on osteogenic marker expression, suggesting that the Ras pathways are necessary but not sufficient for osteogenesis. Taken together, our results indicate that matrix elasticity and cell density are important microenvironmental cues driving hMSCs proliferation and differentiation. Copyright © 2013 Orthopaedic Research Society.

  12. Assembly of fibronectin into the extracellular matrix of early and late passage human skin fibroblasts

    International Nuclear Information System (INIS)

    Mann, D.M.

    1987-01-01

    The specific binding of soluble 125 I-human plasma fibronectin ( 125 I-HFN-P) to confluent cultures of early and late passage human skin fibroblasts was investigated. Previous studies HFN-P bound to fibroblast cell layers indicated that HNF-P was present in the cultures in two separate pools, distinguishable on the basis of their solubility in 1% deoxycholate. Examination of the kinetics of 125 I-HFN-P binding to Pool I of early and late passage cultures revealed that both cultures required 2-4 h to approach steady-state conditions. Other kinetic studies showed that the rates of low of 125 I-HFN-P from either Pool I or Pool II were similar for both cultures. Further, Scatchard analysis revealed a single class of Pool I binding sites with apparent dissociation constants (K/sub d/) of 5.3 x 10 -8 M (early passage) and 4.2 x 10 -8 M (late passage). These results indicate that early and late passage cultures of human fibroblasts exhibit differences in the number of cell surface biding sites for soluble fibronectin, and in the extent to which they incorporate soluble fibronectin into the extracellular matrix. Parameters which affect the fibronectin matrix assembly system of human skin fibroblasts were also examined. In addition, several monoclonal anti-fibronectin antibodies were characterized and developed as experimental probes for fibronectin structure and function

  13. Protein crystallization with microseed matrix screening: application to human germline antibody Fabs

    International Nuclear Information System (INIS)

    Obmolova, Galina; Malia, Thomas J.; Teplyakov, Alexey; Sweet, Raymond W.; Gilliland, Gary L.

    2014-01-01

    The power of microseed matrix screening is demonstrated in the crystallization of a panel of antibody Fab fragments. The crystallization of 16 human antibody Fab fragments constructed from all pairs of four different heavy chains and four different light chains was enabled by employing microseed matrix screening (MMS). In initial screening, diffraction-quality crystals were obtained for only three Fabs, while many Fabs produced hits that required optimization. Application of MMS, using the initial screens and/or refinement screens, resulted in diffraction-quality crystals of these Fabs. Five Fabs that failed to give hits in the initial screen were crystallized by cross-seeding MMS followed by MMS optimization. The crystallization protocols and strategies that resulted in structure determination of all 16 Fabs are presented. These results illustrate the power of MMS and provide a basis for developing future strategies for macromolecular crystallization

  14. Detection of HOCl-mediated protein oxidation products in the extracellular matrix of human atherosclerotic plaques

    DEFF Research Database (Denmark)

    Woods, Alan A; Linton, Stuart M; Davies, Michael Jonathan

    2003-01-01

    Oxidation is believed to play a role in atherosclerosis. Oxidized lipids, sterols and proteins have been detected in early, intermediate and advanced human lesions at elevated levels. The spectrum of oxidized side-chain products detected on proteins from homogenates of advanced human lesions has...... been interpreted in terms of the occurrence of two oxidative mechanisms, one involving oxygen-derived radicals catalysed by trace transition metal ions, and a second involving chlorinating species (HOCl or Cl2), generated by the haem enzyme myeloperoxidase (MPO). As MPO is released extracellularly...... for 83-96% of the total oxidized protein side-chain products detected in these plaques. Oxidation of matrix components extracted from healthy artery tissue, and model proteins, with reagent HOCl is shown to give rise to a similar pattern of products to those detected in advanced human lesions...

  15. Platelet Counts in Insoluble Platelet-Rich Fibrin Clots: A Direct Method for Accurate Determination

    Directory of Open Access Journals (Sweden)

    Yutaka Kitamura

    2018-02-01

    Full Text Available Platelet-rich fibrin (PRF clots have been used in regenerative dentistry most often, with the assumption that growth factor levels are concentrated in proportion to the platelet concentration. Platelet counts in PRF are generally determined indirectly by platelet counting in other liquid fractions. This study shows a method for direct estimation of platelet counts in PRF. To validate this method by determination of the recovery rate, whole-blood samples were obtained with an anticoagulant from healthy donors, and platelet-rich plasma (PRP fractions were clotted with CaCl2 by centrifugation and digested with tissue-plasminogen activator. Platelet counts were estimated before clotting and after digestion using an automatic hemocytometer. The method was then tested on PRF clots. The quality of platelets was examined by scanning electron microscopy and flow cytometry. In PRP-derived fibrin matrices, the recovery rate of platelets and white blood cells was 91.6 and 74.6%, respectively, after 24 h of digestion. In PRF clots associated with small and large red thrombi, platelet counts were 92.6 and 67.2% of the respective total platelet counts. These findings suggest that our direct method is sufficient for estimating the number of platelets trapped in an insoluble fibrin matrix and for determining that platelets are distributed in PRF clots and red thrombi roughly in proportion to their individual volumes. Therefore, we propose this direct digestion method for more accurate estimation of platelet counts in most types of platelet-enriched fibrin matrix.

  16. Decellularized matrix from tumorigenic human mesenchymal stem cells promotes neovascularization with galectin-1 dependent endothelial interaction.

    Directory of Open Access Journals (Sweden)

    Jorge S Burns

    Full Text Available BACKGROUND: Acquisition of a blood supply is fundamental for extensive tumor growth. We recently described vascular heterogeneity in tumours derived from cell clones of a human mesenchymal stem cell (hMSC strain (hMSC-TERT20 immortalized by retroviral vector mediated human telomerase (hTERT gene expression. Histological analysis showed that cells of the most vascularized tumorigenic clone, -BD11 had a pericyte-like alpha smooth muscle actin (ASMA+ and CD146+ positive phenotype. Upon serum withdrawal in culture, -BD11 cells formed cord-like structures mimicking capillary morphogenesis. In contrast, cells of the poorly tumorigenic clone, -BC8 did not stain for ASMA, tumours were less vascularized and serum withdrawal in culture led to cell death. By exploring the heterogeneity in hMSC-TERT20 clones we aimed to understand molecular mechanisms by which mesenchymal stem cells may promote neovascularization. METHODOLOGY/PRINCIPAL FINDINGS: Quantitative qRT-PCR analysis revealed similar mRNA levels for genes encoding the angiogenic cytokines VEGF and Angiopoietin-1 in both clones. However, clone-BD11 produced a denser extracellular matrix that supported stable ex vivo capillary morphogenesis of human endothelial cells and promoted in vivo neovascularization. Proteomic characterization of the -BD11 decellularized matrix identified 50 extracellular angiogenic proteins, including galectin-1. siRNA knock down of galectin-1 expression abrogated the ex vivo interaction between decellularized -BD11 matrix and endothelial cells. More stable shRNA knock down of galectin-1 expression did not prevent -BD11 tumorigenesis, but greatly reduced endothelial migration into -BD11 cell xenografts. CONCLUSIONS: Decellularized hMSC matrix had significant angiogenic potential with at least 50 angiogenic cell surface and extracellular proteins, implicated in attracting endothelial cells, their adhesion and activation to form tubular structures. hMSC -BD11 surface galectin-1

  17. Mesenchymal stromal cells improve human islet function through released products and extracellular matrix.

    Science.gov (United States)

    Arzouni, Ahmed A; Vargas-Seymour, Andreia; Rackham, Chloe L; Dhadda, Paramjeet; Huang, Guo-Cai; Choudhary, Pratik; Nardi, Nance; King, Aileen J F; Jones, Peter M

    2017-12-01

    The aims of the present study were (i) to determine whether the reported beneficial effects of mesenchymal stromal cells (MSCs) on mouse islet function extend to clinically relevant human tissues (islets and MSCs), enabling translation into improved protocols for clinical human islet transplantation; and (ii) to identify possible mechanisms through which human MSCs influence human islet function. Human islets were co-cultured with human adipose tissue-derived MSCs (hASCs) or pre-treated with its products - extracellular matrix (ECM) and annexin A1 (ANXA1). Mouse islets were pre-treated with mouse MSC-derived ECM. Islet insulin secretory function was assessed in vitro by radioimmunoassay. Quantitative RT-PCR was used to screen human adipMSCs for potential ligands of human islet G-protein-coupled receptors. We show that co-culture with hASCs improves human islet secretory function in vitro , as measured by glucose-stimulated insulin secretion, confirming previous reports using rodent tissues. Furthermore, we demonstrate that these beneficial effects on islet function can be partly attributed to the MSC-derived products ECM and ANXA1. Our results suggest that hASCs have the potential to improve the quality of human islets isolated for transplantation therapy of Type 1 diabetes. Furthermore, it may be possible to achieve improvements in human islet quality in a cell-free culture system by using the MSC-derived products ANXA1 and ECM. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  18. Introduction of a potent single-donor fibrin glue for vascular anastomosis: An animal study

    Directory of Open Access Journals (Sweden)

    Mehdi Rasti Ardakani

    2012-01-01

    Full Text Available Background: Vascular anastomosis is considered as a difficult surgical procedure. Although different alternative methods have been tried to tackle these difficulties, none were found to be successful. Commercial fibrin glue has recently been used for vascular anastomosis. However, it did not gain popularity due to some limitations such as low tensile strength, rapid removal by the immune system, and risk of transmission of blood-borne viral infections. In this article, we presented a novel method for producing single-donor human fibrin glue and determined its success rate for vascular anastomosis in an animal model. Materials ans Methods : In this study, 3 mL of single-donor fibrin sealant was prepared from 150 mL of whole blood containing 50-70 mg/mL of fibrinogen. The study was performed on 10 dogs and 5 cats. After transection of the carotid artery, both ends were anastomosed by means of 3-4 sutures (Prolene 8-0. The suture line was then sealed with one layer of the new fibrin sealant. After 3-8 weeks, the site of anastomosis was evaluated angiographically and morphologically for healing and possible complications such as thrombosis or aneurysm. Results: In evaluations 3 weeks after the surgery, all arterial anastomoses were patent in dogs, but some degree of subintimal hyperplasia was noted. After 8 weeks, all anastomoses were patent and the degree of subintimal hyperplasia was decreased. In cats on the other hand, after 4 weeks, all anastomoses were patent and subintimal hyperplasia was absent. Conclusions: Single-donor fibrin glue was a quite reliable and practical alternative to minimize suturing and therefore operative time in our animal model. This sealant can easily be obtained from the patient′s whole blood. Its application in humans would require further studies.

  19. Matrix metalloproteinase sensing via porous silicon microcavity devices functionalized with human antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Marta; Gergely, Csilla [GES-UMR 5650, CNRS, Universite Montpellier 2, Pl. Eugene Bataillon 34095, Montpellier Cedex 5 (France); Taleb Bendiab, Chakib; Massif, Laurent; Cuisinier, Frederic [EA4203, Faculte d' Odontologie, Universite Montpellier 1, Montpellier Cedex 5 (France); Palestino, Gabriela [Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi, Av. Salvador Nava 6, 78000 San Luis Potosi (Mexico); Agarwal, Vivechana [CIICAP, Universidad Autonoma del Estado de Morelos, Av. Universidad 1001, Col Chamilpa, Cuernavaca, Mor. (Mexico)

    2011-06-15

    Porous silicon microcavity (PSiMc) structures were used as support material for specific sensing of matrix metalloproteinases (MMPs). For lower concentrations of MMP-8, the structures were tested with two types of functionalization methods. Silanization of the oxidized porous silicon structures, followed by glutaraldehyde chemistry was found to give very inconsistent results. The use of biotinilated bovine serum albumin linked to the naked PSiMc was found to be an alternative method to attach the anti MMP-8 human antibody, previously modified with streptavidin, which was further used to sense MMP-8 (copyright 2011 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

  20. Role of plasma-rich fibrin in oral surgery

    OpenAIRE

    Kumar, K. Retna; Genmorgan, K.; Abdul Rahman, S. M.; Rajan, M. Alaguvel; Kumar, T. Arul; Prasad, V. Srinivas

    2016-01-01

    Platelet-rich fibrin (PRF) is a fibrin meshwork, in which platelet cytokines, growth factors, and cells are entrapped and discharged after a period and can serve as a resorbable film. PRF is the next generation of platelet concentrates equipped to improve arrangement without biochemical blood handling; PRF is an evolution of the fibrin adhesive, which is widely used in the oral surgery. The guidelines of this innovation depend on concentrating platelets and growth factors in a plasma medium, ...

  1. Engineered matrix coatings to modulate the adhesion of CD133+ human hematopoietic progenitor cells.

    Science.gov (United States)

    Franke, Katja; Pompe, Tilo; Bornhäuser, Martin; Werner, Carsten

    2007-02-01

    Interactions of hematopoietic progenitor cells (HPC) with their local microenvironments in the bone marrow are thought to control homing, differentiation, and self-renewal of the cells. To dissect the role of extracellular matrix (ECM) components of the niche microenvironment, a set of well-defined ECM coatings including fibronectin, heparin, heparan sulphate, hyaluronic acid, tropocollagen I, and co-fibrils of collagen I with heparin or hyaluronic acid was prepared and analysed with respect to the attachment of human CD133+ HPC in vitro. The extension of the adhesion areas of individual cells as well as the fraction of adherent cells were assessed by reflection interference contrast microscopy (RICM). Intense cell-matrix interactions were found on surfaces coated with fibronectin, heparin, heparan sulphate, and on the collagen I based co-fibrils. Insignificant adhesion was found for tropocollagen I and hyaluronic acid. The strongest adhesion of HPC was observed on fibronectin with contact areas of about 7 microm(2). Interaction of HPC with coatings consisting of heparin, heparan sulphate, and co-fibrils result in small circular shaped contact zones of 3 microm(2) pointing to another, less efficient, adhesion mechanism. Analysing the specificity of cell-matrix interaction by antibody blocking experiments suggests an integrin(alpha(5)beta(1))-specific adhesion on fibronectin, while adhesion on heparin was shown to be mediated by selectins (CD62L). Taken together, our data provide a basis for the design of advanced culture carriers supporting site-specific proliferation or differentiation of HPC.

  2. Cytocompatibility and biologic characteristics of synthetic scaffold materials of rabbit acellular vascular matrix combining with human-like collagen I.

    Science.gov (United States)

    Liu, Xuqian; Wang, Jie; Dong, Fusheng; Song, Peng; Tian, Songbo; Li, Hexiang; Hou, Yali

    2017-10-01

    Scaffold material provides a three-dimensional growing environment for seed cells in the research field of tissue engineering. In the present study, rabbit arterial blood vessel cells were chemically removed with trypsin and Triton X-100 to prepare rabbit acellular vascular matrix scaffold material. Observation by He&Masson staining revealed that no cellular components or nuclei existed in the vascular intima and media after decellularization. Human-like collagen I was combined with acellular vascular matrix by freeze-drying to prepare an acellular vascular matrix-0.25% human-like collagen I scaffold to compensate for the extracellular matrix loss during the decellularization process. We next performed a series of experiments to test the water absorbing quality, biomechanics, pressure resistance, cytotoxicity, and ultra-micro structure of the acellular vascular matrix composite material and natural rabbit artery and found that the acellular vascular matrix-0.25% human-like collagen I material behaved similarly to natural rabbit artery. In conclusion, the acellular vascular matrix-0.25% human-like collagen I composite material provides a new approach and lays the foundation for novel scaffold material research into tissue engineering of blood vessels.

  3. Muscarinic receptor agonists stimulate matrix metalloproteinase 1-dependent invasion of human colon cancer cells

    International Nuclear Information System (INIS)

    Raufman, Jean-Pierre; Cheng, Kunrong; Saxena, Neeraj; Chahdi, Ahmed; Belo, Angelica; Khurana, Sandeep; Xie, Guofeng

    2011-01-01

    Highlights: ► Muscarinic receptor agonists stimulated robust human colon cancer cell invasion. ► Anti-matrix metalloproteinase1 antibody pre-treatment blocks cell invasion. ► Bile acids stimulate MMP1 expression, cell migration and MMP1-dependent invasion. -- Abstract: Mammalian matrix metalloproteinases (MMPs) which degrade extracellular matrix facilitate colon cancer cell invasion into the bloodstream and extra-colonic tissues; in particular, MMP1 expression correlates strongly with advanced colon cancer stage, hematogenous metastasis and poor prognosis. Likewise, muscarinic receptor signaling plays an important role in colon cancer; muscarinic receptors are over-expressed in colon cancer compared to normal colon epithelial cells. Muscarinic receptor activation stimulates proliferation, migration and invasion of human colon cancer cells. In mouse intestinal neoplasia models genetic ablation of muscarinic receptors attenuates carcinogenesis. In the present work, we sought to link these observations by showing that MMP1 expression and activation plays a mechanistic role in muscarinic receptor agonist-induced colon cancer cell invasion. We show that acetylcholine, which robustly increases MMP1 expression, stimulates invasion of HT29 and H508 human colon cancer cells into human umbilical vein endothelial cell monolayers – this was abolished by pre-incubation with atropine, a non-selective muscarinic receptor inhibitor, and by pre-incubation with anti-MMP1 neutralizing antibody. Similar results were obtained using a Matrigel chamber assay and deoxycholyltaurine (DCT), an amidated dihydroxy bile acid associated with colon neoplasia in animal models and humans, and previously shown to interact functionally with muscarinic receptors. DCT treatment of human colon cancer cells resulted in time-dependent, 10-fold increased MMP1 expression, and DCT-induced cell invasion was also blocked by pre-treatment with anti-MMP1 antibody. This study contributes to understanding

  4. Muscarinic receptor agonists stimulate matrix metalloproteinase 1-dependent invasion of human colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Raufman, Jean-Pierre, E-mail: jraufman@medicine.umaryland.edu [Division of Gastroenterology and Hepatology, University of Maryland School of Medicine, Baltimore, MD (United States); Cheng, Kunrong; Saxena, Neeraj; Chahdi, Ahmed; Belo, Angelica; Khurana, Sandeep; Xie, Guofeng [Division of Gastroenterology and Hepatology, University of Maryland School of Medicine, Baltimore, MD (United States)

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Muscarinic receptor agonists stimulated robust human colon cancer cell invasion. Black-Right-Pointing-Pointer Anti-matrix metalloproteinase1 antibody pre-treatment blocks cell invasion. Black-Right-Pointing-Pointer Bile acids stimulate MMP1 expression, cell migration and MMP1-dependent invasion. -- Abstract: Mammalian matrix metalloproteinases (MMPs) which degrade extracellular matrix facilitate colon cancer cell invasion into the bloodstream and extra-colonic tissues; in particular, MMP1 expression correlates strongly with advanced colon cancer stage, hematogenous metastasis and poor prognosis. Likewise, muscarinic receptor signaling plays an important role in colon cancer; muscarinic receptors are over-expressed in colon cancer compared to normal colon epithelial cells. Muscarinic receptor activation stimulates proliferation, migration and invasion of human colon cancer cells. In mouse intestinal neoplasia models genetic ablation of muscarinic receptors attenuates carcinogenesis. In the present work, we sought to link these observations by showing that MMP1 expression and activation plays a mechanistic role in muscarinic receptor agonist-induced colon cancer cell invasion. We show that acetylcholine, which robustly increases MMP1 expression, stimulates invasion of HT29 and H508 human colon cancer cells into human umbilical vein endothelial cell monolayers - this was abolished by pre-incubation with atropine, a non-selective muscarinic receptor inhibitor, and by pre-incubation with anti-MMP1 neutralizing antibody. Similar results were obtained using a Matrigel chamber assay and deoxycholyltaurine (DCT), an amidated dihydroxy bile acid associated with colon neoplasia in animal models and humans, and previously shown to interact functionally with muscarinic receptors. DCT treatment of human colon cancer cells resulted in time-dependent, 10-fold increased MMP1 expression, and DCT-induced cell invasion was also blocked by pre

  5. Scaffold architecture and fibrin gels promote meniscal cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Pawelec, K. M., E-mail: pawelec.km@gmail.com, E-mail: jw626@cam.ac.uk; Best, S. M.; Cameron, R. E. [Cambridge Centre for Medical Materials, Materials Science and Metallurgy Department, University of Cambridge, Cambridge CB3 0FS (United Kingdom); Wardale, R. J., E-mail: pawelec.km@gmail.com, E-mail: jw626@cam.ac.uk [Division of Trauma and Orthopaedic Surgery, Department of Surgery, University of Cambridge, Cambridge CB2 2QQ (United Kingdom)

    2015-01-01

    Stability of the knee relies on the meniscus, a complex connective tissue with poor healing ability. Current meniscal tissue engineering is inadequate, as the signals for increasing meniscal cell proliferation have not been established. In this study, collagen scaffold structure, isotropic or aligned, and fibrin gel addition were tested. Metabolic activity was promoted by fibrin addition. Cellular proliferation, however, was significantly increased by both aligned architectures and fibrin addition. None of the constructs impaired collagen type I production or triggered adverse inflammatory responses. It was demonstrated that both fibrin gel addition and optimized scaffold architecture effectively promote meniscal cell proliferation.

  6. Silk-fibrin/hyaluronic acid composite gels for nucleus pulposus tissue regeneration.

    Science.gov (United States)

    Park, Sang-Hyug; Cho, Hongsik; Gil, Eun Seok; Mandal, Biman B; Min, Byoung-Hyun; Kaplan, David L

    2011-12-01

    Scaffold designs are critical for in vitro culture of tissue-engineered cartilage in three-dimensional environments to enhance cellular differentiation for tissue engineering and regenerative medicine. In the present study we demonstrated silk and fibrin/hyaluronic acid (HA) composite gels as scaffolds for nucleus pulposus (NP) cartilage formation, providing both biochemical support for NP outcomes as well as fostering the retention of size of the scaffold during culture due to the combined features of the two proteins. Passage two (P2) human chondrocytes cultured in 10% serum were encapsulated within silk-fibrin/HA gels. Five study groups with fibrin/HA gel culture (F/H) along with varying silk concentrations (2% silk gel only, fibrin/HA gel culture with 1% silk [F/H+1S], 1.5% silk [F/H+1.5S], and 2% silk [F/H+2S]) were cultured in serum-free chondrogenic defined media (CDM) for 4 weeks. Histological examination with alcian blue showed a defined chondrogenic area at 1 week in all groups that widened homogenously until 4 weeks. In particular, chondrogenic differentiation observed in the F/H+1.5S had no reduction in size throughout the culture period. The results of biochemical and molecular biological evaluations supported observations made during histological examination. Mechanical strength measurements showed that the silk mixed gels provided stronger mechanical properties for NP tissue than fibrin/HA composite gels in CDM. This effect could potentially be useful in the study of in vitro NP tissue engineering as well as for clinical implications for NP tissue regeneration.

  7. Demineralized bone matrix and human cancellous bone enhance fixation of titanium implants

    DEFF Research Database (Denmark)

    Babiker, Hassan; Ding, Ming; Overgaard, Søren

    Best Poster 5Demineralized bone matrix and human cancellous bone enhance fixation of titanium implants AuthorsBabiker , H.; Ding M.; Overgaard S.InstitutionOrthopaedic Research Laboratory, Department of Orthopaedic Surgery, Odense University Hospital, Clinical Institute, University of Southern...... from human tissue were included (IsoTis OrthoBiologics, Inc. USA). Both materials are commercially available. Titanium alloy implants (Biomet Inc.) of 10 mm in length and 10 mm in diameter were inserted bilaterally into the femoral condyles of 8 skeletally mature sheep. Thus four implants...... with a concentric gap of 2 mm were implanted in each sheep. The gap was filled with: DBM; DBM/CB with ratio of 1/3; DBM/allograft with ratio of 1/3; or allograft (Gold standard), respectively. Standardised surgical procedure was used1. At sacrifice, 6 weeks after surgery, both distal femurs were harvested...

  8. The Effect of Chondroitin Sulphate and Hyaluronic Acid on Chondrocytes Cultured within a Fibrin-Alginate Hydrogel

    Directory of Open Access Journals (Sweden)

    Christopher J. Little

    2014-09-01

    Full Text Available Osteoarthritis is a painful degenerative joint disease that could be better managed if tissue engineers can develop methods to create long-term engineered articular cartilage tissue substitutes. Many of the tissue engineered cartilage constructs currently available lack the chemical stimuli and cell-friendly environment that promote the matrix accumulation and cell proliferation needed for use in joint cartilage repair. The goal of this research was to test the efficacy of using a fibrin-alginate hydrogel containing hyaluronic acid (HA and/or chondroitin sulphate (CS supplements for chondrocyte culture. Neonatal porcine chondrocytes cultured in fibrin-alginate hydrogels retained their phenotype better than chondrocytes cultured in monolayer, as evidenced by analysis of their relative expression of type II versus type I collagen mRNA transcripts. HA or CS supplementation of the hydrogels increased matrix glycosaminoglycan (GAG production during the first week of culture. However, the effects of these supplements on matrix accumulation were not additive and were no longer observed after two weeks of culture. Supplementation of the hydrogels with CS or a combination of both CS and HA increased the chondrocyte cell population after two weeks of culture. Statistical analysis indicated that the HA and CS treatment effects on chondrocyte numbers may be additive. This research suggests that supplementation with CS and/or HA has positive effects on cartilage matrix production and chondrocyte proliferation in three-dimensional (3D fibrin-alginate hydrogels.

  9. Dynamic imaging in electrical impedance tomography of the human chest with online transition matrix identification.

    Science.gov (United States)

    Moura, Fernando Silva; Aya, Julio Cesar Ceballos; Fleury, Agenor Toledo; Amato, Marcelo Britto Passos; Lima, Raul Gonzalez

    2010-02-01

    One of the electrical impedance tomography objectives is to estimate the electrical resistivity distribution in a domain based only on electrical potential measurements at its boundary generated by an imposed electrical current distribution into the boundary. One of the methods used in dynamic estimation is the Kalman filter. In biomedical applications, the random walk model is frequently used as evolution model and, under this conditions, poor tracking ability of the extended Kalman filter (EKF) is achieved. An analytically developed evolution model is not feasible at this moment. The paper investigates the identification of the evolution model in parallel to the EKF and updating the evolution model with certain periodicity. The evolution model transition matrix is identified using the history of the estimated resistivity distribution obtained by a sensitivity matrix based algorithm and a Newton-Raphson algorithm. To numerically identify the linear evolution model, the Ibrahim time-domain method is used. The investigation is performed by numerical simulations of a domain with time-varying resistivity and by experimental data collected from the boundary of a human chest during normal breathing. The obtained dynamic resistivity values lie within the expected values for the tissues of a human chest. The EKF results suggest that the tracking ability is significantly improved with this approach.

  10. Culture temperature affects redifferentiation and cartilaginous extracellular matrix formation in dedifferentiated human chondrocytes.

    Science.gov (United States)

    Ito, Akira; Aoyama, Tomoki; Iijima, Hirotaka; Tajino, Junichi; Nagai, Momoko; Yamaguchi, Shoki; Zhang, Xiangkai; Kuroki, Hiroshi

    2015-05-01

    To date, there have been few studies on how temperature affects the phenotype and metabolism of human chondrocytes. Thus, the purpose of this study was to elucidate the effects of culture temperature on chondrocyte redifferentiation and extracellular matrix (ECM) formation using dedifferentiated mature human chondrocytes in vitro. Dedifferentiated chondrocytes were cultured in a pellet culture system for up to 21 days. The pellets were randomly divided into three groups with different culture temperature (32, 37, and 41°C). Chondrocyte redifferentiation and ECM formation were evaluated by wet weight, messenger ribonucleic acid (mRNA), histological, and biochemical analyses. The results showed that the wet weight and the mRNA expressions of collagen type II A1 and cartilage oligomeric matrix protein at 37°C were higher than the corresponding values at 32°C. The histological and biochemical analyses revealed that the syntheses of type II collagen and proteoglycan were promoted at 37°C compared to those at 32°C, whereas they were considerably inhibited at 41°C. In conclusion, the results obtained herein indicated that temperature affects chondrocyte redifferentiation and ECM formation, and modulation of temperature might thus represent an advantageous means to regulate the phenotype and biosynthetic activity of chondrocytes. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  11. Experimental closure of gunshot wounds by fibrin glue with antibiotics in pigs

    Directory of Open Access Journals (Sweden)

    Đenić Nebojša

    2015-01-01

    Full Text Available Background/Aim. Gunshot wounds caused by the automatic rifle M70AB2 (AK-47 7.62 mm, after the primary surgical management, were closed with delayed primary suture during the next four to seven days. This period coincides with the fibroblastic phase of wound healing. Fibrin glue is used as a local hemostatic and as a matrix for the local dosed release of antibiotics. Antibiotics addition to fibrin glue resulted in continuous diffusion into the surrounding next 4 to 7 days. The aim of this study was to create the preconditions for gunshot wounds closing without complications by the application of fibrin glue with antibiotics 24 h after primary surgical treatment. Methods. A total of 14 pigs were wounded in the gluteofemoral region by the bullet M67, initial velocity of 720 m/s. All wounded animals were surgically treated according to the principles of the warsurgery doctrine. Seven wounds were closed with primary delayed suture four days after the primary surgical treatment (traditional approach. Fibrin glue with antibiotics was introduced in seven wounds during the primary surgical treatment and primary delayed suture was done after 24 h. The macroscopic appearance and the clinical assessment of the wound were done during the primary surgical treatment and during its revision after 24 h, as well as histopathological findings at the days 4 and 7 after wounding. Results. Gunshot wounds caused by the automatic rifle M70AB2 (AK-47 7.62 mm, and treated with fibrin glue with antibiotics after primary surgical management, were closed with primary delayed suture after 24 h. In further wound evolution there were no complications. Conclusion. Uncomplicated soft-tissue wounds caused by an automatic M70AB2 rifle may be closed primarily with delayed suture without the risk of developing complications if on revision, 24 h after primary surgery, there were no present necrotic tissues, hematoma, and any signs of infection when fibrin glue with antibiotics

  12. HPLC-MS/MS method optimisation for matrix metalloproteinase 3 and matrix metalloproteinase 9 determination in human blood serum using target analysis.

    Science.gov (United States)

    Kotnik, Petra; Krajnc, Metka Koren; Pahor, Artur; Finšgar, Matjaž; Knez, Željko

    2018-02-20

    A quantitative analysis of zinc endopeptidases matrix metalloproteinase 9 (MMP9) and matrix metalloproteinase 3 (MMP3) from human blood serum are presented. Both matrix metalloproteinases (MMP) are present in human blood serum and can be used as biomarkers for different diseases. The analysis was performed using LC-MS/MS with a triple quadrupole mass spectrometer, based on two specific peptides of each MMP in comparison with an enzyme-linked immunosorbent assay (ELISA). While the conditions for the LC-MS/MS analysis of MMP9 peptides were previously reported for bronchoalveolar lavage fluid, the analysis of MMP3 peptides was newly quantified for human blood serum herein for the first time. For MMP3, the linear behaviour was determined in the concentration range from 1.0-200.0ng/mL (R 2 =0.997) with an LLOD of 0.5ng/mL. For MMP9, linearity was determined in the concentration range from 6.5-65.0ng/mL (R 2 =0.995) with an LLOD of 2.0ng/mL. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Oxygen tension is a determinant of the matrix-forming phenotype of cultured human meniscal fibrochondrocytes.

    Directory of Open Access Journals (Sweden)

    Adetola B Adesida

    Full Text Available BACKGROUND: Meniscal cartilage displays a poor repair capacity, especially when injury is located in the avascular region of the tissue. Cell-based tissue engineering strategies to generate functional meniscus substitutes is a promising approach to treat meniscus injuries. Meniscus fibrochondrocytes (MFC can be used in this approach. However, MFC are unable to retain their phenotype when expanded in culture. In this study, we explored the effect of oxygen tension on MFC expansion and on their matrix-forming phenotype. METHODOLOGY/PRINCIPAL FINDINGS: MFC were isolated from human menisci followed by basic fibroblast growth factor (FGF-2 mediated cell expansion in monolayer culture under normoxia (21%O(2 or hypoxia (3%O(2. Normoxia and hypoxia expanded MFC were seeded on to a collagen scaffold. The MFC seeded scaffolds (constructs were cultured in a serum free chondrogenic medium for 3 weeks under normoxia and hypoxia. Constructs containing normoxia-expanded MFC were subsequently cultured under normoxia while those formed from hypoxia-expanded MFC were subsequently cultured under hypoxia. After 3 weeks of in vitro culture, the constructs were assessed biochemically, histologically and for gene expression via real-time reverse transcription-PCR assays. The results showed that constructs under normoxia produced a matrix with enhanced mRNA ratio (3.5-fold higher; p<0.001 of collagen type II to I. This was confirmed by enhanced deposition of collagen II using immuno-histochemistry. Furthermore, the constructs under hypoxia produced a matrix with higher mRNA ratio of aggrecan to versican (3.5-fold, p<0.05. However, both constructs had the same capacity to produce a glycosaminoglycan (GAG -specific extracellular matrix. CONCLUSIONS: Our data provide evidence that oxygen tension is a key player in determining the matrix phenotype of cultured MFC. These findings suggest that the use of normal and low oxygen tension during MFC expansion and subsequent neo

  14. Superoxide produced in the matrix of mitochondria enhances methylmercury toxicity in human neuroblastoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Mailloux, Ryan J.; Yumvihoze, Emmanuel; Chan, Hing Man, E-mail: laurie.chan@uottawa.ca

    2015-12-15

    The mechanism of intracellular metabolism of methylmercury (MeHg) is not fully known. It has been shown that superoxide (O{sub 2}·{sup −}), the proximal reactive oxygen species (ROS) generated by mitochondria, is responsible for MeHg demethylation. Here, we investigated the impact of different mitochondrial respiratory inhibitors, namely rotenone and antimycin A, on the O{sub 2}·{sup −} mediated degradation of MeHg in human neuroblastoma cells SH-K-SN. We also utilized paraquat (PQ) which generates O{sub 2}·{sup −} in the mitochondrial matrix. We found that the cleavage of the carbon-metal bond in MeHg was highly dependent on the topology of O{sub 2}·{sup −} production by mitochondria. Both rotenone and PQ, which increase O{sub 2}·{sup −} in the mitochondrial matrix at a dose-dependent manner, enhanced the conversion of MeHg to inorganic mercury (iHg). Surprisingly, antimycin A, which prompts emission of O{sub 2}·{sup −} into the intermembrane space, did not have the same effect even though antimycin A induced a dose dependent increase in O{sub 2}·{sup −} emission. Rotenone and PQ also enhanced the toxicity of sub-toxic doses (0.1 μM) MeHg which correlated with the accumulation of iHg in mitochondria and depletion of mitochondrial protein thiols. Taken together, our results demonstrate that MeHg degradation is mediated by mitochondrial O{sub 2}·{sup −}, specifically within the matrix of mitochondria when O{sub 2}·{sup −} is in adequate supply. Our results also show that O{sub 2}·{sup −} amplifies MeHg toxicity specifically through its conversion to iHg and subsequent interaction with protein cysteine thiols (R-SH). The implications of our findings in mercury neurotoxicity are discussed herein. - Highlights: • Superoxide produced in the matrix of mitochondria degrades MeHg. • Superoxide produced in intermembrane space does not degrade MeHg. • Matrix-generated superoxide enhances Hg toxicity by converting MeHg to iHg.

  15. Contaminating fibrin in CPD-blood: solubility in plasma and distribution in blood components following separation

    International Nuclear Information System (INIS)

    Skjonsberg, O.H.; Kierulf, P.; Gravem, K.; Fagerhol, M.K.; Godal, H.C.

    1986-01-01

    In order to estimate the solubility of contaminating fibrin in CPD-blood, thrombin induced fibrin polymerzation in CPD-plasma was examined by light scattering and fibrinopeptide A (FPA) determinations. In addition, I-125 fibrin monomer enriched CPD-blood was used to investigate fibrin monomer retention in blood bags and transfusion filters (170 microns) and fibrin distribution in blood components derived from CPD-blood. Initial fibrin polymerization in CPD-blood occurred after conversion of 15 per cent of the fibrinogen to fibrin, implying that substantial amounts of fibrin may be kept solubilized in CPD-blood bags. Only minor amounts of I-125 fibrin monomers were retained in blood bags (2.4 per cent) and in transfusion filters (2.9 per cent) after sham transfusions. After separating I-125-fibrin monomer enriched CPD-blood into its constituent components, the major part of fibrin (75.0 per cent) could be traced in the cryoprecipitate

  16. USAGE OF MONOCLONAL ANTIBODIES FOR DETERMINATION OF LOCALIZATION OF ANTIGENIC DETERMINANTS AND FIBRIN POLYMERIZATION SITES WITHIN FIBRINOGEN AND FIBRIN MOLECULES AND THEIR APPLICATION IN TEST--SYSTEMS FOR DIAGNOSTICS AND THE THREAT OF THROMBUS FORMATION

    Directory of Open Access Journals (Sweden)

    E. V. Lugovskoi

    2013-08-01

    Full Text Available It was shown by monoclonal antibodies that B?N-region of fibrin desA molecule (B?1-53 comprises the polymerization site including the peptide bond B?14-15. This site participates in the second stage of fibrin polymerization — lateral association of protofibrils. In the B?15-53 fragment was also found the site called «C», which together with the site «A» participate in the first stage of polymerization — the protofibrils formation. The model of the primary intermolecular interaction of fibrin was designed. It was found by monoclonal antibodies II-4d the site («c» in the N-terminal half of ? chain of the fibrin D-region. This site participates in the protofibrils formation and is complement to site «C» as we assume. We have discovered two neoantigenic determinants. One of these determinants exposes within the coiledcoil fragment B?126-135 of fibrin as a result of fibrinopeptide A splitting off from fibrinogen by thrombin. The structural rearrangements discovered in this site of the fibrin molecule are necessary for the following protofibrils lateral association. The second neoantigenic determinant is localized in the fragment B?134-190 of D-dimer formed after plasmin degradation of fibrin stabilized by FXIIIa. We have obtained the fibrin-specific monoclonal antibodie FnI-3C to the first determinant and D-dimer-specific mAb III-3b to the second one. Three monoclonal antibodies were obtained against the ?C-region of fibrin(ogen molecule. It has been experimentally shown by of one of them that ?C-domains is connected with the fibrinopeptides B in fibrinogen and fibrin desA molecules, but removes from the core of the molecules after fibrinopeptides B splitting off by thrombin. Two other monoclonal antibodies specifically inhibit the fibrin polymerization by blocking two unknown polymerization sites within the ?C-region. The test-systems for the soluble fibrin and D-dimer quantification in human blood plasma were designed on the basis of

  17. Engineering fibrin hydrogels to promote the wound healing potential of mesenchymal stem cell spheroids.

    Science.gov (United States)

    Murphy, Kaitlin C; Whitehead, Jacklyn; Zhou, Dejie; Ho, Steve S; Leach, J Kent

    2017-12-01

    Mesenchymal stem cells (MSCs) secrete endogenous factors such as vascular endothelial growth factor (VEGF) and prostaglandin E2 (PGE 2 ) that promote angiogenesis, modulate the inflammatory microenvironment, and stimulate wound repair, and MSC spheroids secrete more trophic factors than dissociated, individual MSCs. Compared to injection of cells alone, transplantation of MSCs in a biomaterial can enhance their wound healing potential by localizing cells at the defect site and upregulating trophic factor secretion. To capitalize on the therapeutic potential of spheroids, we engineered a fibrin gel delivery vehicle to simultaneously enhance the proangiogenic and anti-inflammatory potential of entrapped human MSC spheroids. We used multifactorial statistical analysis to determine the interaction between four input variables derived from fibrin gel synthesis on four output variables (gel stiffness, gel contraction, and secretion of VEGF and PGE 2 ). Manipulation of the four input variables tuned fibrin gel biophysical properties to promote the simultaneous secretion of VEGF and PGE 2 by entrapped MSC spheroids while maintaining overall gel integrity. MSC spheroids in stiffer gels secreted the most VEGF, while PGE 2 secretion was highest in more compliant gels. Simultaneous VEGF and PGE 2 secretion was greatest using hydrogels with intermediate mechanical properties, as small increases in stiffness increased VEGF secretion while maintaining PGE 2 secretion by entrapped spheroids. The fibrin gel formulation predicted to simultaneously increase VEGF and PGE 2 secretion stimulated endothelial cell proliferation, enhanced macrophage polarization, and promoted angiogenesis when used to treat a wounded three-dimensional human skin equivalent. These data demonstrate that a statistical approach is an effective strategy to formulate fibrin gel formulations that enhance the wound healing potential of human MSCs. Mesenchymal stem cells (MSCs) are under investigation for wound

  18. Human Adipose Tissue Derived Extracellular Matrix and Methylcellulose Hydrogels Augments and Regenerates the Paralyzed Vocal Fold.

    Directory of Open Access Journals (Sweden)

    Dong Wook Kim

    Full Text Available Vocal fold paralysis results from various etiologies and can induce voice changes, swallowing complications, and issues with aspiration. Vocal fold paralysis is typically managed using injection laryngoplasty with fat or synthetic polymers. Injection with autologous fat has shown excellent biocompatibility. However, it has several disadvantages such as unpredictable resorption rate, morbidities associated with liposuction procedure which has to be done in operating room under general anesthesia. Human adipose-derived extracellular matrix (ECM grafts have been reported to form new adipose tissue and have greater biostability than autologous fat graft. Here, we present an injectable hydrogel that is constructed from adipose tissue derived soluble extracellular matrix (sECM and methylcellulose (MC for use in vocal fold augmentation. Human sECM derived from adipose tissue was extracted using two major steps-ECM was isolated from human adipose tissue and was subsequently solubilized. Injectable sECM/MC hydrogels were prepared by blending of sECM and MC. Sustained vocal fold augmentation and symmetric vocal fold vibration were accomplished by the sECM/MC hydrogel in paralyzed vocal fold which were confirmed by laryngoscope, histology and a high-speed imaging system. There were increased number of collagen fibers and fatty granules at the injection site without significant inflammation or fibrosis. Overall, these results indicate that the sECM/MC hydrogel can enhance vocal function in paralyzed vocal folds without early resorption and has potential as a promising material for injection laryngoplasty for stable vocal fold augmentation which can overcome the shortcomings of autologous fat such as unpredictable duration and morbidity associated with the fat harvest.

  19. Human Adipose Tissue Derived Extracellular Matrix and Methylcellulose Hydrogels Augments and Regenerates the Paralyzed Vocal Fold.

    Science.gov (United States)

    Kim, Dong Wook; Kim, Eun Ji; Kim, Eun Na; Sung, Myung Whun; Kwon, Tack-Kyun; Cho, Yong Woo; Kwon, Seong Keun

    2016-01-01

    Vocal fold paralysis results from various etiologies and can induce voice changes, swallowing complications, and issues with aspiration. Vocal fold paralysis is typically managed using injection laryngoplasty with fat or synthetic polymers. Injection with autologous fat has shown excellent biocompatibility. However, it has several disadvantages such as unpredictable resorption rate, morbidities associated with liposuction procedure which has to be done in operating room under general anesthesia. Human adipose-derived extracellular matrix (ECM) grafts have been reported to form new adipose tissue and have greater biostability than autologous fat graft. Here, we present an injectable hydrogel that is constructed from adipose tissue derived soluble extracellular matrix (sECM) and methylcellulose (MC) for use in vocal fold augmentation. Human sECM derived from adipose tissue was extracted using two major steps-ECM was isolated from human adipose tissue and was subsequently solubilized. Injectable sECM/MC hydrogels were prepared by blending of sECM and MC. Sustained vocal fold augmentation and symmetric vocal fold vibration were accomplished by the sECM/MC hydrogel in paralyzed vocal fold which were confirmed by laryngoscope, histology and a high-speed imaging system. There were increased number of collagen fibers and fatty granules at the injection site without significant inflammation or fibrosis. Overall, these results indicate that the sECM/MC hydrogel can enhance vocal function in paralyzed vocal folds without early resorption and has potential as a promising material for injection laryngoplasty for stable vocal fold augmentation which can overcome the shortcomings of autologous fat such as unpredictable duration and morbidity associated with the fat harvest.

  20. Effect of Human Serum and 2 Different Types of Platelet Concentrates on Human Meniscus Cell Migration, Proliferation, and Matrix Formation.

    Science.gov (United States)

    Freymann, Undine; Metzlaff, Sebastian; Krüger, Jan-Philipp; Hirsh, Glen; Endres, Michaela; Petersen, Wolf; Kaps, Christian

    2016-06-01

    To evaluate the effect of 10% human serum (HS), 5% platelet-rich plasma (PRP), and 5% autologous conditioned plasma (ACP) on migration, proliferation, and extracellular matrix (ECM) synthesis of human meniscus cells. Cell migration and proliferation on stimulation with HS, PRP, and ACP were assessed by chemotaxis assays and measurement of genomic DNA content. Meniscus cells were cultivated in pellets stimulated with 10% HS, 5% PRP, or 5% ACP. Meniscal ECM formation was evaluated by histochemical staining of collagen type I, type II, and proteoglycans and by analysis of fibrochondrocyte marker gene expression. Human meniscus cells were significantly attracted by all 3 blood-derived products (10% HS and 5% ACP: P = .0001, 5% PRP: P = .0002). Cell proliferation at day 9 was significantly increased on stimulation with 10% HS (P = .0001) and 5% PRP (P = .0002) compared with 5% ACP and controls. Meniscus cell pellet cultures showed the formation of a well-structured meniscal ECM with deposition of collagen type I, type II, and proteoglycans on stimulation with 10% HS, whereas 5% PRP or 5% ACP resulted in the formation of an inhomogeneous and more fibrous ECM. Stimulation with 10% HS and 5% ACP showed a significant induction of fibrochondrocyte marker genes such as aggrecan (HS: P = .0002, ACP: P = .0147), cartilage oligomeric matrix protein (HS: P = .0002, ACP: P = .0005), and biglycan (HS: P = .0002, ACP: P = .0003), whereas PRP showed no inducing effect. Among all tested blood-derived products, only stimulation with HS showed the formation of a meniscal ECM as well as positive cell proliferating and migrating effects in vitro. Regarding a potential biological repair of nonvascular meniscus lesions, our results may point toward the use of HS as a beneficial augment in regenerative meniscus repair approaches. Our findings may suggest that HS might be a beneficial augment for meniscus repair. Copyright © 2016 Arthroscopy Association of North America. Published

  1. A novel fibrin gel derived from hyaluronic acid-grafted fibrinogen

    International Nuclear Information System (INIS)

    Yang, Chiung L; Chen, Hui W; Wang, Tzu C; Wang, Yng J

    2011-01-01

    Fibrinogen is a major plasma protein that forms a three-dimensional fibrin gel upon being activated by thrombin. In this study, we report the synthesis and potential applications of hybrid molecules composed of fibrinogen coupled to the reducing ends of short-chain hyaluronic acids (sHAs) by reductive amination. The grafting of sHAs to fibrinogen was verified by analyzing particle size, zeta potential and gel-electrophoretic mobility of the hybrid molecules. The sHA-fibrinogen hybrid molecules with graft ratios (sHA/fibrinogen) of up to 6.5 retained the ability to form gels in response to thrombin activation. The sHA-fibrin gels were transparent in appearance and exhibited high water content, which were characteristics distinct from those of gels formed by mixtures of sHAs and fibrinogen. The potential applications of the sHA-fibrin gels were evaluated. The sHA-fibrinogen gel with a graft ratio of 3.6 (S3.6F) was examined for its ability to encapsulate and support the differentiation of ATDC5 chondrocyte-like cells. Compared with the fibrinogen-formed gel, cells cultured in the S3.6F gel exhibited increased lacunae formation; moreover, the abundance of cartilaginous extracellular matrix molecules and the expression of chondrocyte marker genes, such as aggrecan, collagen II and Sox9, were also significantly increased. Our data suggest that the three-dimensional gel formed by the sHA-fibrinogen hybrid is a better support than the fibrin gel for chondrogenesis induction.

  2. Scintigraphic evaluation of osteoblastic activity in extraction sockets treated with platelet-rich fibrin.

    Science.gov (United States)

    Gürbüzer, Bahadir; Pikdöken, Levent; Tunali, Mustafa; Urhan, Muammer; Küçükodaci, Zafer; Ercan, Feriha

    2010-05-01

    To evaluate the effect of platelet-rich fibrin (PRF) on the early bone healing process with bone scintigraphy based on technetium-99m methylene diphosphonate uptake in third molar extraction sockets. Fourteen patients with bilaterally soft tissue impacted third mandibular molars were included in the study. The right and left impacted third molars were surgically extracted in the same session. PRF was randomly administered into one of the extraction sockets, whereas the contralateral sockets were left without treatment. Four weeks after surgery, scintigrams were obtained to evaluate scintigraphic differences between PRF-treated and non-PRF-treated sockets. After completion of the clinical study, PRF samples were evaluated by light and scanning electron microscopy. The average increase in technetium-99m methylene diphosphonate uptake as an indication of enhanced bone healing did not differ significantly between PRF-treated and non-PRF-treated sockets 4 weeks postoperatively (P > .05). Abundant fibrin and inflammatory cells were observed by light microscopic examination of PRF samples. Scanning electron microscopic analysis of PRF revealed the existence of platelet aggregates in a fibrin network and crystalline particles on the outer surface of PRF. PRF might not lead to enhanced bone healing in soft tissue impacted mandibular third molar extraction sockets 4 weeks after surgery. PRF exhibits the potential characteristics of an autologous fibrin matrix. However, whether the presence of crystal-like particles on the outer surface of PRF alters bone healing should be investigated further. Copyright 2010 American Association of Oral and Maxillofacial Surgeons. Published by Elsevier Inc. All rights reserved.

  3. A novel fibrin gel derived from hyaluronic acid-grafted fibrinogen

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Chiung L; Chen, Hui W; Wang, Tzu C; Wang, Yng J, E-mail: wang@ym.edu.tw [Institute of Biomedical Engineering, National Yang Ming University, No. 155, Sec. 2, Li-Nung St., Shih-Pai, Taipei, Taiwan 112 (China)

    2011-04-15

    Fibrinogen is a major plasma protein that forms a three-dimensional fibrin gel upon being activated by thrombin. In this study, we report the synthesis and potential applications of hybrid molecules composed of fibrinogen coupled to the reducing ends of short-chain hyaluronic acids (sHAs) by reductive amination. The grafting of sHAs to fibrinogen was verified by analyzing particle size, zeta potential and gel-electrophoretic mobility of the hybrid molecules. The sHA-fibrinogen hybrid molecules with graft ratios (sHA/fibrinogen) of up to 6.5 retained the ability to form gels in response to thrombin activation. The sHA-fibrin gels were transparent in appearance and exhibited high water content, which were characteristics distinct from those of gels formed by mixtures of sHAs and fibrinogen. The potential applications of the sHA-fibrin gels were evaluated. The sHA-fibrinogen gel with a graft ratio of 3.6 (S3.6F) was examined for its ability to encapsulate and support the differentiation of ATDC5 chondrocyte-like cells. Compared with the fibrinogen-formed gel, cells cultured in the S3.6F gel exhibited increased lacunae formation; moreover, the abundance of cartilaginous extracellular matrix molecules and the expression of chondrocyte marker genes, such as aggrecan, collagen II and Sox9, were also significantly increased. Our data suggest that the three-dimensional gel formed by the sHA-fibrinogen hybrid is a better support than the fibrin gel for chondrogenesis induction.

  4. Delineation of in vitro chondrogenesis of human synovial stem cells following preconditioning using decellularized matrix

    Science.gov (United States)

    Zhang, Ying; Li, Jingting; Davis, Mary E.; Pei, Ming

    2015-01-01

    As a tissue-specific stem cell for chondrogenesis, synovium-derived stem cells (SDSCs) are a promising cell source for cartilage repair. However, a small biopsy can only provide a limited number of cells. Cell senescence from both in vitro expansion and donor age presents a big challenge for stem cell based cartilage regeneration. Here we found that expansion on decellularized extracellular matrix (dECM) full of three-dimensional nanostructured fibers provided SDSCs with unique surface profiles, low elasticity but large volume as well as fibroblast-like shape. dECM expanded SDSCs yielded larger pellets with intensive staining of type II collagen and sulfated glycosaminoglycans compared to those grown on plastic flasks while SDSCs grown in ECM yielded 28-day pellets with minimal matrix as evidenced by pellet size and chondrogenic marker staining, which was confirmed by both biochemical data and real-time PCR data. Our results also found lower levels of inflammatory genes in dECM expanded SDSCs that might be responsible for enhanced chondrogenic differentiation. Despite an increase in type X collagen in chondrogenically induced cells, dECM expanded cells had significantly lower potential for endochondral bone formation. Wnt and MAPK signals were actively involved in both expansion and chondrogenic induction of dECM expanded cells. Since young and healthy people can be potential donors for this matrix expansion system and decellularization can minimize immune concerns, human SDSCs expanded on this future commercially available dECM could be a potential cell source for autologous cartilage repair. PMID:25861949

  5. Use of fibrin sealants in cardiovascular surgery: a systematic review.

    Science.gov (United States)

    Rousou, John A

    2013-05-01

    Fibrin sealants are used for hemostasis and tissue adherence. This systematic review summarizes published clinical data for fibrin sealant use in cardiovascular surgery. A literature search for the following terms was conducted using PubMed and EMBASE: (TISSEEL or Tissucol or Beriplast P or Evicel or Quixil or Crosseal or Reliseal or Fibringluraas or Bolheal or Tachosil or Vivostat or Vitagel or Artiss or "fibrin glue" or "fibrin sealant" or "fibrin tissue adhesive") and (cardiac or cardiovascular or vascular or heart or coronary or surgery). Case reports and series were excluded; although reports of controlled trials were preferred, uncontrolled trial data were also considered. Clinical trials and chart review analyses of fibrin sealants were identified and summarized. Although clinical trial data were available for other agents, the majority of published studies examined TISSEEL. Overall, TISSEEL and other fibrin sealants showed improvements over standard of care or control groups for a variety of predefined endpoints. Safety findings are also summarized. Data from these studies showed that fibrin sealants were well tolerated and provided effective hemostasis in a range of cardiac and aortic surgeries. © 2013 Wiley Periodicals, Inc.

  6. Circulating Microparticles Alter Formation, Structure, and Properties of Fibrin Clots.

    Science.gov (United States)

    Zubairova, Laily D; Nabiullina, Roza M; Nagaswami, Chandrasekaran; Zuev, Yuriy F; Mustafin, Ilshat G; Litvinov, Rustem I; Weisel, John W

    2015-12-04

    Despite the importance of circulating microparticles in haemostasis and thrombosis, there is limited evidence for potential causative effects of naturally produced cell-derived microparticles on fibrin clot formation and its properties. We studied the significance of blood microparticles for fibrin formation, structure, and susceptibility to fibrinolysis by removing them from platelet-free plasma using filtration. Clots made in platelet-free and microparticle-depleted plasma samples from the same healthy donors were analyzed in parallel. Microparticles accelerate fibrin polymerisation and support formation of more compact clots that resist internal and external fibrinolysis. These variations correlate with faster thrombin generation, suggesting thrombin-mediated kinetic effects of microparticles on fibrin formation, structure, and properties. In addition, clots formed in the presence of microparticles, unlike clots from the microparticle-depleted plasma, contain 0.1-0.5-μm size granular and CD61-positive material on fibres, suggesting that platelet-derived microparticles attach to fibrin. Therefore, the blood of healthy individuals contains functional microparticles at the levels that have a procoagulant potential. They affect the structure and stability of fibrin clots indirectly through acceleration of thrombin generation and through direct physical incorporation into the fibrin network. Both mechanisms underlie a potential role of microparticles in haemostasis and thrombosis as modulators of fibrin formation, structure, and resistance to fibrinolysis.

  7. Host-Parasite Interaction: Parasite-Derived and -Induced Proteases That Degrade Human Extracellular Matrix

    Directory of Open Access Journals (Sweden)

    Carolina Piña-Vázquez

    2012-01-01

    Full Text Available Parasitic protozoa are among the most important pathogens worldwide. Diseases such as malaria, leishmaniasis, amoebiasis, giardiasis, trichomoniasis, and trypanosomiasis affect millions of people. Humans are constantly threatened by infections caused by these pathogens. Parasites engage a plethora of surface and secreted molecules to attach to and enter mammalian cells. The secretion of lytic enzymes by parasites into host organs mediates critical interactions because of the invasion and destruction of interstitial tissues, enabling parasite migration to other sites within the hosts. Extracellular matrix is a complex, cross-linked structure that holds cells together in an organized assembly and that forms the basement membrane lining (basal lamina. The extracellular matrix represents a major barrier to parasites. Therefore, the evolution of mechanisms for connective-tissue degradation may be of great importance for parasite survival. Recent advances have been achieved in our understanding of the biochemistry and molecular biology of proteases from parasitic protozoa. The focus of this paper is to discuss the role of protozoan parasitic proteases in the degradation of host ECM proteins and the participation of these molecules as virulence factors. We divide the paper into two sections, extracellular and intracellular protozoa.

  8. Grooved surface topography alters matrix-metalloproteinase production by human fibroblasts

    International Nuclear Information System (INIS)

    Brydone, Alistair S; Dominic Meek, R M; Dalby, Matthew J; Berry, Catherine C; McNamara, Laura E

    2011-01-01

    Extracellular matrix (ECM) remodelling is an essential physiological process in which matrix-metalloproteinases (MMPs) have a key role. Manipulating the manner in which cells produce MMPs and ECMs may enable the creation of a desired tissue type, i.e. effect repair, or the prevention of tissue invasion (e.g. metastasis). The aim of this project was to determine if culturing fibroblasts on grooved topography altered collagen deposition or MMP production. Human fibroblasts were seeded on planar or grooved polycaprolactone substrates (grooves were 12.5 μm wide with varying depths of 240 nm, 540 nm or 2300 nm). Cell behaviour and collagen production were studied using fluorescence microscopy and the spent culture medium was assessed using gel zymography to detect MMPs. Total collagen deposition was high on the 240 nm deep grooves, but decreased as the groove depth increased, i.e. as cell contact guidance decreased. There was an increase in gelatinase on the 2300 nm deep grooved topography and there was a difference in the temporal expression of MMP-3 observed on the planar surface compared to the 540 nm and 2300 nm topographies. These results show that topography can alter collagen and MMP production. A fuller understanding of these processes may permit the design of surfaces tailored to tissue regeneration e.g. tendon repair.

  9. Food Matrix Effects of Polyphenol Bioaccessibility from Almond Skin during Simulated Human Digestion

    Directory of Open Access Journals (Sweden)

    Giuseppina Mandalari

    2016-09-01

    Full Text Available The goal of the present study was to quantify the rate and extent of polyphenols released in the gastrointestinal tract (GIT from natural (NS and blanched (BS almond skins. A dynamic gastric model of digestion which provides a realistic simulation of the human stomach was used. In order to establish the effect of a food matrix on polyphenols bioaccessibility, NS and BS were either digested in water (WT or incorporated into home-made biscuits (HB, crisp-bread (CB and full-fat milk (FM. Phenolic acids were the most bioaccessible class (68.5% release from NS and 64.7% from BS. WT increased the release of flavan-3-ols (p < 0.05 and flavonols (p < 0.05 from NS after gastric plus duodenal digestion, whereas CB and HB were better vehicles for BS. FM lowered the % recovery of polyphenols, the free total phenols and the antioxidant status in the digestion medium, indicating that phenolic compounds could bind protein present in the food matrix. The release of bioactives from almond skins could explain the beneficial effects associated with almond consumption.

  10. Buddleja officinalis inhibits high glucose-induced matrix metalloproteinase activity in human umbilical vein endothelial cells.

    Science.gov (United States)

    Lee, Yun Jung; Kang, Dae Gill; Kim, Jin Sook; Lee, Ho Sub

    2008-12-01

    The aim of the present investigation was to investigate whether an aqueous extract of Buddleja officinalis (ABO), a traditional Korean herbal medicine, suppresses the endothelial extracellular matrix degradation under high glucose condition. The incubation with high concentration of glucose (25 mM) increased significantly matrix metalloproteinase (MMP)-2/-9 expressions and activities in primary cultured human umbilical vein endothelial cells (HUVEC). Pretreatment with ABO decreased high glucose-induced increase of MMP-2/-9 activities in a dose-dependent manner. Real time qRT-PCR revealed that high glucose-induced MMP-2/-9 mRNA expression levels were attenuated by pretreatment with ABO. High glucose-induced MCP-1 and IL-8 mRNA expression levels also decreased by ABO. ABO decreased high glucose-induced hydrogen peroxide production, oxidative stress marker. These results provide new insights into the pathophysiological mechanisms for anti-inflammatory properties of ABO in vascular diseases associated with diabetes mellitus. (c) 2008 John Wiley & Sons, Ltd.

  11. Survival of Lactobacillus rhamnosus strains inoculated in cheese matrix during simulated human digestion.

    Science.gov (United States)

    Pitino, Iole; Randazzo, Cinzia L; Cross, Kathryn L; Parker, Mary L; Bisignano, Carlo; Wickham, Martin S J; Mandalari, Giuseppina; Caggia, Cinzia

    2012-08-01

    Survival of probiotic bacteria during transit through the gastrointestinal (GI) tract is influenced by a number of environmental variables including stomach acidity, bile salts, digestive enzymes and food matrix. This study assessed survival of seven selected Lactobacillus rhamnosus strains delivered within a model cheese system to the human upper GI tract using a dynamic gastric model (DGM). Good survival rates for all tested strains were recorded during both simulated gastric and duodenal digestion. Strains H12, H25 and N24 demonstrated higher survival capacities during gastric digestion than L. rhamnosus GG strain used as control, with H12 and N24 continuing to grow during duodenal digestion. Strains L. rhamnosus F17, N24 and R61 showed adhesion properties to both HT-29 and Caco-2 cells. The ability to attach to the cheese matrix during digestion was confirmed by scanning electron microscopy, also indicating production of extracellular polysaccharides as a response to acid stress. Copyright © 2012 Elsevier Ltd. All rights reserved.

  12. Grooved surface topography alters matrix-metalloproteinase production by human fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Brydone, Alistair S; Dominic Meek, R M [Department of Orthopaedics, Southern General Hospital, 1345 Govan Road, Glasgow G51 4TF (United Kingdom); Dalby, Matthew J; Berry, Catherine C; McNamara, Laura E, E-mail: alibrydone@gmail.com [Centre for Cell Engineering, Joseph Black Building, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ (United Kingdom)

    2011-06-15

    Extracellular matrix (ECM) remodelling is an essential physiological process in which matrix-metalloproteinases (MMPs) have a key role. Manipulating the manner in which cells produce MMPs and ECMs may enable the creation of a desired tissue type, i.e. effect repair, or the prevention of tissue invasion (e.g. metastasis). The aim of this project was to determine if culturing fibroblasts on grooved topography altered collagen deposition or MMP production. Human fibroblasts were seeded on planar or grooved polycaprolactone substrates (grooves were 12.5 {mu}m wide with varying depths of 240 nm, 540 nm or 2300 nm). Cell behaviour and collagen production were studied using fluorescence microscopy and the spent culture medium was assessed using gel zymography to detect MMPs. Total collagen deposition was high on the 240 nm deep grooves, but decreased as the groove depth increased, i.e. as cell contact guidance decreased. There was an increase in gelatinase on the 2300 nm deep grooved topography and there was a difference in the temporal expression of MMP-3 observed on the planar surface compared to the 540 nm and 2300 nm topographies. These results show that topography can alter collagen and MMP production. A fuller understanding of these processes may permit the design of surfaces tailored to tissue regeneration e.g. tendon repair.

  13. Food Matrix Effects of Polyphenol Bioaccessibility from Almond Skin during Simulated Human Digestion

    Science.gov (United States)

    Mandalari, Giuseppina; Vardakou, Maria; Faulks, Richard; Bisignano, Carlo; Martorana, Maria; Smeriglio, Antonella; Trombetta, Domenico

    2016-01-01

    The goal of the present study was to quantify the rate and extent of polyphenols released in the gastrointestinal tract (GIT) from natural (NS) and blanched (BS) almond skins. A dynamic gastric model of digestion which provides a realistic simulation of the human stomach was used. In order to establish the effect of a food matrix on polyphenols bioaccessibility, NS and BS were either digested in water (WT) or incorporated into home-made biscuits (HB), crisp-bread (CB) and full-fat milk (FM). Phenolic acids were the most bioaccessible class (68.5% release from NS and 64.7% from BS). WT increased the release of flavan-3-ols (p digestion, whereas CB and HB were better vehicles for BS. FM lowered the % recovery of polyphenols, the free total phenols and the antioxidant status in the digestion medium, indicating that phenolic compounds could bind protein present in the food matrix. The release of bioactives from almond skins could explain the beneficial effects associated with almond consumption. PMID:27649239

  14. Alterations in the extracellular matrix organization associated with the reexpression of tumorigenicity in human cell hybrids.

    Science.gov (United States)

    Der, C J; Stanbridge, E J

    1980-10-15

    The expression of fibronectin on the cell surface was evaluated on a series of intraspecific human cell hybrids formed between HeLa and normal fibroblast strains. Although these hybrids continued to express many of the in vitro transformation properties of their corresponding tumorigenic HeLa parent, they were now unable to form tumors when inoculated into athymic nude mice. From these suppressed hybrid populations, rare tumorigenic segregant subpopulations arose which had regained their tumorigenic capacity. A comparison of the expression of fibronectin on the cell surface was made between these tumorigenic segregant cell lines and their corresponding non-tumorigenic HeLa/fibroblast hybrid. Following specific immunofluorescent staining for fibronectin, a striking alteration in the cell surface organization was observed to correspond with the reexpression of tumorigenicity in these hybrids. Tumorigenic HeLa/fibroblast hybrids were also significantly altered in both their cellular and colonial morphology. Double immunofluorescent staining to simultaneously visualize both surface fibronectin and collagen revealed that these two extracellular matrix proteins displayed an extensive degree of codistribution and expressed a coordinate shift in organization which correlated with the appearance of tumorigenic segregant hybrid populations. These observations are in agreement with the apparently close structural association between fibronectin and collagen and suggest that the organization of these two components in the extracellular matrix may be an important determinant for in vivo growth potential.

  15. Growth factor and proteinase profile of Vivostat® platelet-rich fibrin linked to tissue repair.

    Science.gov (United States)

    Agren, M S; Rasmussen, K; Pakkenberg, B; Jørgensen, B

    2014-07-01

    Autologous platelet-rich fibrin (PRF(®)) is prepared by the automatic Vivostat(®) system. Conflicting results with Vivostat PRF in acute wound healing prompted us to examine its cellular and biomolecular composition. Specifically, platelets, selected growth factors and matrix metalloproteinase (MMP)-9 were quantified using novel analytical methods. Ten healthy non-thrombocytopenic volunteers donated blood for generation of intermediate fibrin-I and final PRF. Anticoagulated whole blood and serum procured in parallel served as baseline controls. Leucocyte, erythrocyte and platelet counts in whole blood and fibrin-I were determined by automated haematology analyser. Platelet concentration in PRF was quantified manually by stereologic analysis of Giemsa-stained tissue sections, and the total content of five growth factors and MMP-9 by enzyme-linked immunosorbent assays. The number of leucocytes and erythrocytes was reduced (P platelets increased (P fibrin-I versus whole blood. PRF contained 982 ± 206 × 10(9) platelets/l representing 3·9-fold (P platelet-derived growth factor (PDGF)-AB [2·5-fold, P PDGF-BB [1·6-fold, P vascular endothelial growth factor > basic fibroblast growth factor [75-fold, P platelet enrichment and biomolecular constituents may guide clinicians in their optimal use of Vivostat PRF for tissue regenerative applications. © 2013 International Society of Blood Transfusion.

  16. Spontaneous and cytokine induced expression and activity of matrix metalloproteinases in human colonic epithelium

    DEFF Research Database (Denmark)

    Pedersen, G; Saermark, T; Kirkegaard, T

    2009-01-01

    levels in cells from inflamed IBD mucosa. MMP-2 and -8 mRNA were expressed inconsistently and MMP-11, -13 and -14 mRNA undetectable. Proteolytic MMP activity was detected in CEC supernatants and the level was increased significantly in inflamed IBD epithelium. The enzyme activity was inhibited strongly......Matrix metalloproteinases (MMPs) have been implicated in tissue damage associated with inflammatory bowel disease (IBD).As the role of the intestinal epithelium in this process is unknown, we determined MMP expression and enzyme activity in human colonic epithelial cells (CEC). MMP mRNA expression...... was assessed by reverse transcription-polymerase chain reaction in HT-29 and DLD-1 cells and in CEC isolated from biopsies from IBD and control patients. Total MMP activity in the cells was measured by a functional assay, based on degradation of a fluorescent synthetic peptide containing the specific bond...

  17. Expression of matrix metrallproteinase-2 in human tears fluid after LASIK

    Directory of Open Access Journals (Sweden)

    Ai-Wei Chen

    2014-12-01

    Full Text Available AIM: To monitor long-term changes of matrix metalloproteinase-2(MMP-2in human tears fluid after laser in situ keratomileusis(LASIK. METHODS: Thirty-two myopia cases(64 eyesunderwent uneventful LASIK were enrolled in the study. Tear fluid were collected and MMP-2 expression was analyzed by Western-bolt assay preoperatively and postoperatively on 15d, at 1, 3mo, and 1a. RESULTS: LASIK increased the concentration of MMP-2 in human tear fluid. At 15d postoperatively, the magnitude of MMP-2 was 1.4 times that of preoperative, thereafter subsided, but didn't return to preoperative level by 3mo(PP>0.05. CONCLUSION: MMP-2 is significantly expressed in human tear fluid after LASIK, then subsided with time, but didn't return to preoperative level by 3mo and almost recovered up to 1a, indicating wound healing of LASIK would continue up at least 3mo after surgery and almost recovered 1a postoperatively.

  18. Comparative Proteomic Analysis of Supportive and Unsupportive Extracellular Matrix Substrates for Human Embryonic Stem Cell Maintenance*

    Science.gov (United States)

    Soteriou, Despina; Iskender, Banu; Byron, Adam; Humphries, Jonathan D.; Borg-Bartolo, Simon; Haddock, Marie-Claire; Baxter, Melissa A.; Knight, David; Humphries, Martin J.; Kimber, Susan J.

    2013-01-01

    Human embryonic stem cells (hESCs) are pluripotent cells that have indefinite replicative potential and the ability to differentiate into derivatives of all three germ layers. hESCs are conventionally grown on mitotically inactivated mouse embryonic fibroblasts (MEFs) or feeder cells of human origin. In addition, feeder-free culture systems can be used to support hESCs, in which the adhesive substrate plays a key role in the regulation of stem cell self-renewal or differentiation. Extracellular matrix (ECM) components define the microenvironment of the niche for many types of stem cells, but their role in the maintenance of hESCs remains poorly understood. We used a proteomic approach to characterize in detail the composition and interaction networks of ECMs that support the growth of self-renewing hESCs. Whereas many ECM components were produced by supportive and unsupportive MEF and human placental stromal fibroblast feeder cells, some proteins were only expressed in supportive ECM, suggestive of a role in the maintenance of pluripotency. We show that identified candidate molecules can support attachment and self-renewal of hESCs alone (fibrillin-1) or in combination with fibronectin (perlecan, fibulin-2), in the absence of feeder cells. Together, these data highlight the importance of specific ECM interactions in the regulation of hESC phenotype and provide a resource for future studies of hESC self-renewal. PMID:23658023

  19. A Practical Method ‘Discussion using Matrix Diagram’ , ConnectingHuman Base-Liberal-and Engineering Base-Professional-

    Science.gov (United States)

    Shimada, Wataru

    In order to bring up talented people, it is a most important subject how to awake ‘Emotional Human Power’ , which is the origin of Autonomy and Creativity. A Practical Method ‘Discussion using Matrix Diagram’ developed for improving ‘Emotional Human Power’ including ‘Communication Skill’ , is confirmed to be useful for connecting Human Base-Liberal-and Engineering Base-Professional-.

  20. Interface between breast cancer cells and the tumor microenvironment using platelet-rich plasma to promote tumor angiogenesis - influence of platelets and fibrin bundles on the behavior of breast tumor cells.

    Science.gov (United States)

    Andrade, Sheila Siqueira; Sumikawa, Joana Tomomi; Castro, Eloísa Dognani; Batista, Fabricio Pereira; Paredes-Gamero, Edgar; Oliveira, Lilian Carolina; Guerra, Izabel Monastério; Peres, Giovani Bravin; Cavalheiro, Renan Pelluzzi; Juliano, Luiz; Nazário, Afonso Pinto; Facina, Gil; Tsai, Siu Mui; Oliva, Maria Luiza Vilela; Girão, Manoel João Batista Castello

    2017-03-07

    Cancer progression is associated with an evolving tissue interface of direct epithelial-tumor microenvironment interactions. In biopsies of human breast tumors, extensive alterations in molecular pathways are correlated with cancer staging on both sides of the tumor-stroma interface. These interactions provide a pivotal paracrine signaling to induce malignant phenotype transition, the epithelial-mesenchymal transition (EMT). We explored how the direct contact between platelets-fibrin bundles primes metastasis using platelet-rich plasma (PRP) as a source of growth factors and mimics the provisional fibrin matrix between actively growing breast cancer cells and the tumor stroma. We have demonstrated PRP functions, modulating cell proliferation that is tumor-subtype and cancer cell-type-specific. Epithelial and stromal primary cells were prepared from breast cancer biopsies from 21 women with different cancer subtypes. Cells supplemented with PRP were immunoblotted with anti-phospho and total Src-Tyr-416, FAK-Try-925, E-cadherin, N-cadherin, TGF-β, Smad2, and Snail monoclonal antibodies. Breast tumor cells from luminal B and HER2 subtypes showed the most malignant profiles and the expression of thrombin and other classes of proteases at levels that were detectable through FRET peptide libraries. The angiogenesis process was investigated in the interface obtained between platelet-fibrin-breast tumor cells co-cultured with HUVEC cells. Luminal B and HER2 cells showed robust endothelial cell capillary-like tubes ex vivo. The studied interface contributes to the attachment of endothelial cells, provides a source of growth factors, and is a solid substrate. Thus, replacement of FBS supplementation with PRP supplementation represents an efficient and simple approach for mimicking the real multifactorial tumor microenvironment.

  1. Identification of transcriptional macromolecular associations in human bone using browser based in silico analysis in a giant correlation matrix.

    Science.gov (United States)

    Reppe, Sjur; Sachse, Daniel; Olstad, Ole K; Gautvik, Vigdis T; Sanderson, Paul; Datta, Harish K; Berg, Jens P; Gautvik, Kaare M

    2013-03-01

    Intracellular signaling is critically dependent on gene regulatory networks comprising physical molecular interactions. Presently, there is a lack of comprehensive databases for most human tissue types to verify such macromolecular interactions. We present a user friendly browser which helps to identify functional macromolecular interactions in human bone as significant correlations at the transcriptional level. The molecular skeletal phenotype has been characterized by transcriptome analysis of iliac crest bone biopsies from 84 postmenopausal women through quantifications of ~23,000 mRNA species. When the signal levels were inter-correlated, an array containing >260 million correlations was generated, thus recognizing the human bone interactome at the RNA level. The matrix correlation and p values were made easily accessible by a freely available online browser. We show that significant correlations within the giant matrix are reproduced in a replica set of 13 male vertebral biopsies. The identified correlations differ somewhat from transcriptional interactions identified in cell culture experiments and transgenic mice, thus demonstrating that care should be taken in extrapolating such results to the in vivo situation in human bone. The current giant matrix and web browser are a valuable tool for easy access to the human bone transcriptome and molecular interactions represented as significant correlations at the RNA-level. The browser and matrix should be a valuable hypothesis generating tool for identification of regulatory mechanisms and serve as a library of transcript relationships in human bone, a relatively inaccessible tissue. Copyright © 2012 Elsevier Inc. All rights reserved.

  2. Retina tissue engineering by conjunctiva mesenchymal stem cells encapsulated in fibrin gel: Hypotheses on novel approach to retinal diseases treatment.

    Science.gov (United States)

    Soleimannejad, Mostafa; Ebrahimi-Barough, Somayeh; Nadri, Samad; Riazi-Esfahani, Mohammad; Soleimani, Masoud; Tavangar, Seyed Mohammad; Ai, Jafar

    2017-04-01

    Retinitis pigmentosa (RP) and age related macular degeneration (AMD) are two retinal diseases that progress by photoreceptor cells death. In retinal transplantation studies, stem and progenitor cells inject into the sub retinal space or vitreous and then these cells can be migrate to the site of retinal degeneration and locate in the host retina and restitute vision. Our hypothesis suggests that using human conjunctiva stem cells (as the source for increasing the number of human stem cells progenitor cells in retina dysfunction diseases) with fibrin gel and also assessing its relating in vitro (cellular and molecular processes) and in vivo (vision tests and pathology) could be a promising strategy for treatment of AMD and RP disorders. In this idea, we describe a novel approach for retina tissue engineering with differentiation of conjunctiva mesenchymal stem cells (CJMSCs) into photoreceptor-like cells in fibrin gel with induction medium contain taurine. For assessment of differentiation, immunocytochemistry and real time PCR are used for the expression of Rhodopsin, RPE65, Nestin as differentiated photoreceptor cell markers in 2D and 3D culture. The results show that fibrin gel will offer a proper 3D scaffold for CJMSCs derived photoreceptor cell-like cells. Application of immune-privileged, readily available sources of adult stem cells like human conjunctiva stem cells with fibrin gel would be a promising strategy to increase the number of photoreceptor progenitor cells and promote involuntary angiogenesis needed in retina layer repair and regeneration. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. The fast release of stem cells from alginate-fibrin microbeads in injectable scaffolds for bone tissue engineering

    Science.gov (United States)

    Zhou, Hongzhi; Xu, Hockin H. K.

    2011-01-01

    Stem cell-encapsulating hydrogel microbeads of several hundred microns in size suitable for injection, that could quickly degrade to release the cells, are currently unavailable. The objectives of this study were to: (1) develop oxidized alginate-fibrin microbeads encapsulating human umbilical cord mesenchymal stem cells (hUCMSCs); (2) investigate microbead degradation, cell release, and osteogenic differentiation of the released cells for the first time. Three types of microbeads were fabricated to encapsulate hUCMSCs: (1) Alginate microbeads; (2) oxidized alginate microbeads; (3) oxidized alginate-fibrin microbeads. Microbeads with sizes of about 100–500 µm were fabricated with 1×106 hUCMSCs/mL of alginate. For the alginate group, there was little microbead degradation, with very few cells released at 21 d. For oxidized alginate, the microbeads started to slightly degrade at 14 d. In contrast, the oxidized alginate-fibrin microbeads started to degrade at 4 d and released the cells. At 7 d, the number of released cells greatly increased and showed a healthy polygonal morphology. At 21 d, the oxidized alginate-fibrin group had a live cell density that was 4-fold that of the oxidized alginate group, and 15-fold that of the alginate group. The released cells had osteodifferentiation, exhibiting highly elevated bone marker gene expressions of ALP, OC, collagen I, and Runx2. Alizarin staining confirmed the synthesis of bone minerals by hUCMSCs, with the mineral concentration at 21 d being 10-fold that at 7 d. In conclusion, fast-degradable alginate-fibrin microbeads with hUCMSC encapsulation were developed that could start to degrade and release the cells at 4 d. The released hUCMSCs had excellent proliferation, osteodifferentiation, and bone mineral synthesis. The alginate-fibrin microbeads are promising to deliver stem cells inside injectable scaffolds to promote tissue regeneration. PMID:21757229

  4. Wound healing and degradation of the fibrin sealant Beriplast P following partial liver resection in rabbits.

    Science.gov (United States)

    Kroez, Monika; Lang, Wiegand; Dickneite, Gerhard

    2005-01-01

    The objective of this study was to investigate the degradation kinetics of the fibrin sealant (FS) Beriplast P in an experimental liver surgery model in rabbits. A partial liver resection was performed in 21 rabbits, and the wound area covered with Beriplast P to ensure hemostasis. Wound healing of the resection sites was evaluated morphologically over 11 weeks. Degradation of the FS was evaluated by measuring the thickness of the remaining fibrin layer. Plasma samples were analyzed for antibodies against fibrinogen, albumin, thrombin, fibrin, and factor XIII. No postoperative hemorrhage was observed, indicating successful hemostasis throughout. The FS was degraded with a half-life of about 25 days postapplication and was completely replaced by granulation tissue within 9 weeks. The FS degradation and tissue development followed the general stages of wound healing: inflammation and resorption, proliferation, organization and production of collagen, maturation, and scarring. An immune reaction was elicited against the main four human proteins of the FS. The antibody titers peaked on day 14, with a gradual decrease thereafter. We conclude that the FS accomplished hemostasis, facilitated healing in accordance with natural processes, and was completely degraded over time. In humans, the reduced immunogenicity of the FS would potentially increase its degradation half-life.

  5. Equibiaxial cyclic stretch stimulates fibroblasts to rapidly remodel fibrin.

    Science.gov (United States)

    Balestrini, Jenna Leigh; Billiar, Kristen Lawrence

    2006-01-01

    Understanding the effects of the mechanical environment on wound healing is critical for developing more effective treatments to reduce scar formation and contracture. The aim of this study was to investigate the effects of dynamic mechanical stretch on cell-mediated early wound remodeling independent of matrix alignment which obscures more subtle remodeling mechanisms. Cyclic equibiaxial stretch (16% stretch at 0.2 Hz) was applied to fibroblast-populated fibrin gel in vitro wound models for eight days. Compaction, density, tensile strength, and collagen content were quantified as functional measures of remodeling. Stretched samples were approximately ten times stronger, eight-fold more dense, and eight times thinner than statically cultured samples. These changes were accompanied by a 15% increase in net collagen but no significant differences in cell number or viability. When collagen crosslinking was inhibited in stretched samples, the extensibility increased and the strength decreased. The apparent weakening was due to a reduction in compaction rather than a decrease in ability of the tissue to withstand tensile forces. Interestingly, inhibiting collagen crosslinking had no measurable effects on the statically cultured samples. These results indicate that amplified cell-mediated compaction and even a slight addition in collagen content play substantial roles in mechanically induced wound strengthening. These findings increase our understanding of how mechanical forces guide the healing response in skin, and the methods employed in this study may also prove valuable tools for investigating stretch-induced remodeling of other planar connective tissues and for creating mechanically robust engineered tissues.

  6. Predictive factors for anterior chamber fibrin formation after vitreoretinal surgery

    Directory of Open Access Journals (Sweden)

    Leonardo Provetti Cunha

    2014-04-01

    Full Text Available Purpose: The aim of this study was to investigate possible predictive factors related to anterior chamber fibrin formation after vitreoretinal surgery in a large series of patients. Methods: The data of 185 eyes of 185 patients submitted to vitreoretinal surgery was reviewed. The following variables were evaluated: the postoperatively presence of fibrin, age, diabetes mellitus, the vitrectomy system gauge (20, 23 or 25 gauge, the type of vitreous substitute, the influence of prior surgical procedures and the combination with cataract extraction. To evaluate predictive factors for anterior chamber fibrin formation, univariate analysis was performed. A multivariate stepwise logistic regression model was adjusted to investigate factors associated with fibrin formation (p<0.05. Results: Fibrinoid anterior chamber reaction was found in 12 (6.4% patients. For multivariate logistic regression analysis, balanced salt solution (BSS, the chance of fibrin occurrence was 5 times greater (odds ratio 4.83, CI 95% 1.302 - 17.892; p=0.019, while combination with phacoemulsification increased the chance of fibrin formation by 20 times (odds ratio 20, CI 95% 2.480 - 161.347; p=0.005. No significant difference was found regarding other variables. Conclusion: Anterior chamber fibrin formation is an unwanted complication after vitreoretinal surgery. Factors such as combined performance of phacoemulsification and the use of balanced salt solution as a vitreous substitute may predispose the occurrence of this complication.

  7. Role of plasma-rich fibrin in oral surgery

    Directory of Open Access Journals (Sweden)

    K Retna Kumar

    2016-01-01

    Full Text Available Platelet-rich fibrin (PRF is a fibrin meshwork, in which platelet cytokines, growth factors, and cells are entrapped and discharged after a period and can serve as a resorbable film. PRF is the next generation of platelet concentrates equipped to improve arrangement without biochemical blood handling; PRF is an evolution of the fibrin adhesive, which is widely used in the oral surgery. The guidelines of this innovation depend on concentrating platelets and growth factors in a plasma medium, and initiating them in a fibrin gel, keeping in mind the end goal to enhance the healing of wounds. Maxillary bone loss requires numerous regenerative techniques: as a supplement to the procedures of tissue regeneration, a platelet concentrate called PRF was tested for the 1st time in France by Dr. Choukroun. This article enriches the benefits and role of plasma-rich fibrin in oral surgery. Platelet-concentrate fibrin is an evolution of the fibrin glue, which is widely used in the oral surgery.

  8. Chondrogenesis of human infrapatellar fat pad stem cells on acellular dermal matrix

    Directory of Open Access Journals (Sweden)

    Ken eYe

    2016-01-01

    Full Text Available Acellular dermal matrix (ADM has been in clinical use for decades in numerous surgical applications. The ability for ADM to promote cellular repopulation and revascularisation, and tissue regeneration is well documented. Adipose stem cells have the ability to differentiate into mesenchymal tissue types, including bone and cartilage. The aim of this study was to investigate the potential interaction between ADM and adipose stem cells in vitro using TGFβ3 and BMP6.Human infrapatellar fat pad derived adipose stem cells (IPFP-ASC were cultured with ADM derived from rat dermis under chondrogenic (TGFβ3 and BMP6 in vitro for 2 and 4 weeks. Histology, qPCR and immunohistochemistry were performed to assess for markers of chondrogenesis (collagen Type II, SOX9 and proteoglycans. At 4 weeks, cell-scaffold constructs displayed cellular changes consistent with chondrogenesis, with evidence of stratification of cell layers and development of a hyaline-like cartilage layer superficially which stained positively for collagen Type II and proteoglycans. Significant cell-matrix interaction was seen between the cartilage layer and the ADM itself with seamless integration between each layer. Real time qPCR showed significantly increases of COL2A1, SOX9, and ACAN gene expression over 4 weeks when compared to control. COL1A2 gene expression remained unchanged over 4 weeks.We believe the principles which make ADM versatile and successful for tissue regeneration are application to cartilage regeneration. This study demonstrates in vitro the ability for IPFP-ASCs to undergo chondrogenesis, infiltrate and interact with ADM. These outcomes serve as a platform for in vivo modelling of ADM for cartilage repair.

  9. Expression pattern of matrix metalloproteinases in human gynecological cancer cell lines

    International Nuclear Information System (INIS)

    Schröpfer, Andrea; Kammerer, Ulrike; Kapp, Michaela; Dietl, Johannes; Feix, Sonja; Anacker, Jelena

    2010-01-01

    Matrix metalloproteinases (MMPs) are involved in the degradation of protein components of the extracellular matrix and thus play an important role in tumor invasion and metastasis. Their expression is related to the progression of gynecological cancers (e.g. endometrial, cervical or ovarian carcinoma). In this study we investigated the expression pattern of the 23 MMPs, currently known in humans, in different gynecological cancer cell lines. In total, cell lines from three endometrium carcinomas (Ishikawa, HEC-1-A, AN3 CA), three cervical carcinomas (HeLa, Caski, SiHa), three chorioncarcinomas (JEG, JAR, BeWo), two ovarian cancers (BG-1, OAW-42) and one teratocarcinoma (PA-1) were examined. The expression of MMPs was analyzed by RT-PCR, Western blot and gelatin zymography. We demonstrated that the cell lines examined can constitutively express a wide variety of MMPs on mRNA and protein level. While MMP-2, -11, -14 and -24 were widely expressed, no expression was seen for MMP-12, -16, -20, -25, -26, -27 in any of the cell lines. A broad range of 16 MMPs could be found in the PA1 cells and thus this cell line could be used as a positive control for general MMP experiments. While the three cervical cancer cell lines expressed 10-14 different MMPs, the median expression in endometrial and choriocarcinoma cells was 7 different enzymes. The two investigated ovarian cancer cell lines showed a distinctive difference in the number of expressed MMPs (2 vs. 10). Ishikawa, Caski, OAW-42 and BeWo cell lines could be the best choice for all future experiments on MMP regulation and their role in endometrial, cervical, ovarian or choriocarcinoma development, whereas the teratocarcinoma cell line PA1 could be used as a positive control for general MMP experiments

  10. Response of induced bone defects in horses to collagen matrix containing the human parathyroid hormone gene.

    Science.gov (United States)

    Backstrom, Kristin C; Bertone, Alicia L; Wisner, Erik R; Weisbrode, Stephen E

    2004-09-01

    To determine whether human parathyroid hormone (hPTH) gene in collagen matrix could safely promote bone formation in diaphyseal or subchondral bones of horses. 8 clinically normal adult horses. Amount, rate, and quality of bone healing for 13 weeks were determined by use of radiography, quantitative computed tomography, and histomorphometric analysis. Diaphyseal cortex and subchondral bone defects of metacarpi were filled with hPTH(1-34) gene-activated matrix (GAM) or remained untreated. Joints were assessed on the basis of circumference, synovial fluid analysis, pain on flexion, lameness, and gross and histologic examination. Bone volume index was greater for cortical defects treated with hPTH(1-34) GAM, compared with untreated defects. Bone production in cortical defects treated with hPTH(1-34) GAM positively correlated with native bone formation in untreated defects. In contrast, less bone was detected in hPTH(1-34) GAM-treated subchondral bone defects, compared with untreated defects, and histology confirmed poorer healing and residual collagen sponge. Use of hPTH(1-34) GAM induced greater total bone, specifically periosteal bone, after 13 weeks of healing in cortical defects of horses. The hPTH(1-34) GAM impeded healing of subchondral bone but was biocompatible with joint tissues. Promotion of periosteal bone formation may be beneficial for healing of cortical fractures in horses, but the delay in onset of bone formation may negate benefits. The hPTH(1-34) GAM used in this study should not be placed in articular subchondral bone defects, but contact with articular surfaces is unlikely to cause short-term adverse effects.

  11. The human metapneumovirus matrix protein stimulates the inflammatory immune response in vitro.

    Directory of Open Access Journals (Sweden)

    Audrey Bagnaud-Baule

    Full Text Available Each year, during winter months, human Metapneumovirus (hMPV is associated with epidemics of bronchiolitis resulting in the hospitalization of many infants. Bronchiolitis is an acute illness of the lower respiratory tract with a consequent inflammation of the bronchioles. The rapid onset of inflammation suggests the innate immune response may have a role to play in the pathogenesis of this hMPV infection. Since, the matrix protein is one of the most abundant proteins in the Paramyxoviridae family virion, we hypothesized that the inflammatory modulation observed in hMPV infected patients may be partly associated with the matrix protein (M-hMPV response. By western blot analysis, we detected a soluble form of M-hMPV released from hMPV infected cell as well as from M-hMPV transfected HEK 293T cells suggesting that M-hMPV may be directly in contact with antigen presenting cells (APCs during the course of infection. Moreover, flow cytometry and confocal microscopy allowed determining that M-hMPV was taken up by dendritic cells (moDCs and macrophages inducing their activation. Furthermore, these moDCs enter into a maturation process inducing the secretion of a broad range of inflammatory cytokines when exposed to M-hMPV. Additionally, M-hMPV activated DCs were shown to stimulate IL-2 and IFN-γ production by allogeneic T lymphocytes. This M-hMPV-mediated activation and antigen presentation of APCs may in part explain the marked inflammatory immune response observed in pathology induced by hMPV in patients.

  12. Autologous fibrin glue as an encapsulating scaffold for delivery of retinal progenitor cells

    Directory of Open Access Journals (Sweden)

    Tamer Anwar Esmail Ahmed

    2015-02-01

    Full Text Available The retina is a highly sophisticated piece of the neural machinery that begins the translation of incoming light signals into meaningful visual information. Several degenerative diseases of the retina are characterized by photoreceptor loss and eventually lead to irreversible blindness. Regenerative medicine, using tissue engineering-based constructs to deliver progenitor cells or photoreceptors along with supporting carrier matrix is a promising approach for restoration of structure and function. Fresh fibrin glue (FG produced by the CryoSeal®FS system in combination with mouse retinal progenitor cells (RPCs were evaluated in this study. In vitro expanded RPCs isolated from postnatal mouse retina were encapsulated into FG and cultured in the presence of the protease inhibitor, tranexamic acid. Encapsulation of RPCs into FG did not show adverse effects on cell proliferation or cell survival. RPCs exhibited fibroblast-like morphology concomitantly with attachment to the encapsulating FG surface. They expressed α7 and β3 integrin subunits that could mediate attachment to fibrin matrix via an RGD independent mechanism. The three dimensional environment and the attachment surface provided by FG was associated with a rapid downregulation of the progenitor marker SOX2 and enhanced the expression of the differentiation markers CRX and recoverin. However, the in vitro culture conditions did not promote full differentiation into mature photoreceptors. Nevertheless, we have shown that autologous fibrin, when fabricated into a scaffold for RPCs for delivery to the retina, provides the cells with external cues that could potentially improve the differentiation events. Hence, transient encapsulation of RPCs into FG could be a valid and potential treatment strategy to promote retinal regeneration following degenerative diseases. However, further optimization is necessary to maximize the outcomes in terms of mature photoreceptors.

  13. CADM1 controls actin cytoskeleton assembly and regulates extracellular matrix adhesion in human mast cells.

    Directory of Open Access Journals (Sweden)

    Elena P Moiseeva

    Full Text Available CADM1 is a major receptor for the adhesion of mast cells (MCs to fibroblasts, human airway smooth muscle cells (HASMCs and neurons. It also regulates E-cadherin and alpha6beta4 integrin in other cell types. Here we investigated a role for CADM1 in MC adhesion to both cells and extracellular matrix (ECM. Downregulation of CADM1 in the human MC line HMC-1 resulted not only in reduced adhesion to HASMCs, but also reduced adhesion to their ECM. Time-course studies in the presence of EDTA to inhibit integrins demonstrated that CADM1 provided fast initial adhesion to HASMCs and assisted with slower adhesion to ECM. CADM1 downregulation, but not antibody-dependent CADM1 inhibition, reduced MC adhesion to ECM, suggesting indirect regulation of ECM adhesion. To investigate potential mechanisms, phosphotyrosine signalling and polymerisation of actin filaments, essential for integrin-mediated adhesion, were examined. Modulation of CADM1 expression positively correlated with surface KIT levels and polymerisation of cortical F-actin in HMC-1 cells. It also influenced phosphotyrosine signalling and KIT tyrosine autophosphorylation. CADM1 accounted for 46% of surface KIT levels and 31% of F-actin in HMC-1 cells. CADM1 downregulation resulted in elongation of cortical actin filaments in both HMC-1 cells and human lung MCs and increased cell rigidity of HMC-1 cells. Collectively these data suggest that CADM1 is a key adhesion receptor, which regulates MC net adhesion, both directly through CADM1-dependent adhesion, and indirectly through the regulation of other adhesion receptors. The latter is likely to occur via docking of KIT and polymerisation of cortical F-actin. Here we propose a stepwise model of adhesion with CADM1 as a driving force for net MC adhesion.

  14. Reticuloendothelial hyperphagocytosis occurs in streptozotocin-diabetic rats. Studies with colloidal carbon, albumin microaggregates, and soluble fibrin monomers

    International Nuclear Information System (INIS)

    Cornell, R.P.

    1982-01-01

    In contrast to previous studies of diabetic humans and animals, which reported unchanged or depressed function, reticuloendothelial system (RES) hyperphagocytosis of colloidal carbon, 125 I-albumin microaggregates, and 125 I-fibrin monomers were observed in rats as early as 14 days after the induction of diabetes with streptozotocin (STZ). The fact that enhanced phagocytosis by RE macrophages was prevented by chronic insulin replacement therapy indicates that the diabetic internal environment of hyperglycemia and hypoinsulinemia was perhaps responsible for the observed changes. Experiments involving organ localization of intravenously administered particles, perfusion of isolated livers, and microscopic examination of the liver all suggested that increased Kupffer cell activity was the primary event in RES hyperphagocytosis by STZ-diabetic rats. Both hypertrophy and hyperplasia of Kupffer cells were apparent in livers of STZ-diabetic animals as evidenced by photomicrographs and hepatic cell quantification. Plasma fibronectin, which binds fibrin monomers to RE macrophages before phagocytosis, was significantly decreased in the circulation of STZ-diabetic rats, but the level of cell-associated fibronectin was not measured. Renal localization of urea-soluble 125 I-fibrin monomers exceeded splenic and pulmonary uptake in normal control rats and was enhanced in animals with STZ-diabetes. Changes in fibronectin levels, fibrin monomer localization, and Kupffer cell size and numbers in experimental diabetes in rats may have implications for the pathogenesis of vascular disease involving phagocytic mesangial and foam cells in diabetic humans

  15. Reticuloendothelial hyperphagocytosis occurs in streptozotocin-diabetic rats. Studies with colloidal carbon, albumin microaggregates, and soluble fibrin monomers.

    Science.gov (United States)

    Cornell, R P

    1982-02-01

    In contrast to previous studies of diabetic humans and animals, which reported unchanged or depressed function, reticuloendothelial system (RES) hyperphagocytosis of colloidal carbon, 125I-albumin microaggregates, and 125I-fibrin monomers were observed in rats as early as 14 days after the induction of diabetes with streptozotocin (STZ). The fact that enhanced phagocytosis by RE macrophages was prevented by chronic insulin replacement therapy indicates that the diabetic internal environment of hyperglycemia and hypoinsulinemia was perhaps responsible for the observed changes. Experiments involving organ localization of intravenously administered particles, perfusion of isolated livers, and microscopic examination of the liver all suggested that increased Kupffer cell activity was the primary event in RES hyperphagocytosis by STZ-diabetic rats. Both hypertrophy and hyperplasia of Kupffer cells were apparent in livers of STZ-diabetic animals as evidenced by photomicrographs and hepatic cell quantification. Plasma fibronectin, which binds fibrin monomers to RE macrophages before phagocytosis, was significantly decreased in the circulation of STZ-diabetic rats, but the level of cell-associated fibronectin was not measured. Renal localization of urea-soluble 125I-fibrin monomers exceeded splenic and pulmonary uptake in normal control rats and was enhanced in animals with STZ-diabetes. Changes in fibronectin levels, fibrin monomer localization, and Kupffer cell size and numbers in experimental diabetes in rats may have implications for the pathogenesis of vascular disease involving phagocytic mesangial and foam cells in diabetic humans.

  16. Extracellular matrix production by human osteoblasts cultured on biodegradable polymers applicable for tissue engineering.

    Science.gov (United States)

    El-Amin, S F; Lu, H H; Khan, Y; Burems, J; Mitchell, J; Tuan, R S; Laurencin, C T

    2003-03-01

    The nature of the extracellular matrix (ECM) is crucial in regulating cell functions via cell-matrix interactions, cytoskeletal organization, and integrin-mediated signaling. In bone, the ECM is composed of proteins such as collagen (CO), fibronectin (FN), laminin (LM), vitronectin (VN), osteopontin (OP) and osteonectin (ON). For bone tissue engineering, the ECM should also be considered in terms of its function in mediating cell adhesion to biomaterials. This study examined ECM production, cytoskeletal organization, and adhesion of primary human osteoblastic cells on biodegradable matrices applicable for tissue engineering, namely polylactic-co-glycolic acid 50:50 (PLAGA) and polylactic acid (PLA). We hypothesized that the osteocompatible, biodegradable polymer surfaces promote the production of bone-specific ECM proteins in a manner dependent on polymer composition. We first examined whether the PLAGA and PLA matrices could support human osteoblastic cell growth by measuring cell adhesion at 3, 6 and 12h post-plating. Adhesion on PLAGA was consistently higher than on PLA throughout the duration of the experiment, and comparable to tissue culture polystyrene (TCPS). ECM components, including CO, FN, LM, ON, OP and VN, produced on the surface of the polymers were quantified by ELISA and localized by immunofluorescence staining. All of these proteins were present at significantly higher levels on PLAGA compared to PLA or TCPS surfaces. On PLAGA, OP and ON were the most abundant ECM components, followed by CO, FN, VN and LN. Immunofluorescence revealed an extracellular distribution for CO and FN, whereas OP and ON were found both intracellularly as well as extracellularly on the polymer. In addition, the actin cytoskeletal network was more extensive in osteoblasts cultured on PLAGA than on PLA or TCPS. In summary, we found that osteoblasts plated on PLAGA adhered better to the substrate, produced higher levels of ECM molecules, and showed greater cytoskeletal

  17. Tumour imaging by the detection of fibrin clots in tumour stroma using an anti-fibrin Fab fragment

    Science.gov (United States)

    Obonai, Toshifumi; Fuchigami, Hirobumi; Furuya, Fumiaki; Kozuka, Naoyuki; Yasunaga, Masahiro; Matsumura, Yasuhiro

    2016-01-01

    The diagnosis of early and aggressive types of cancer is important for providing effective cancer therapy. Cancer-induced fibrin clots exist only within lesions. Previously, we developed a monoclonal antibody (clone 102-10) that recognizes insoluble fibrin but not fibrinogen or soluble fibrin and confirmed that fibrin clots form continuously in various cancers. Here, we describe the development of a Fab fragment probe of clone 102-10 for tumour imaging. The distribution of 102-10 Fab was investigated in genetically engineered mice bearing pancreatic ductal adenocarcinoma (PDAC), and its effect on blood coagulation was examined. Immunohistochemical and ex vivo imaging revealed that 102-10 Fab was distributed selectively in fibrin clots in PDAC tumours 3 h after injection and that it disappeared from the body after 24 h. 102-10 Fab had no influence on blood coagulation or fibrinolysis. Tumour imaging using anti-fibrin Fab may provide a safe and effective method for the diagnosis of invasive cancers by detecting fibrin clots in tumour stroma. PMID:27009516

  18. Investigating the role of the extracellular matrix on differentiation of human mesenchymal stem cells and MC3T3 cells

    NARCIS (Netherlands)

    Fernandes, H.A.M.; Dechering, Koen; Someren, Eugene; van Blitterswijk, Clemens; de Boer, Jan

    2008-01-01

    Human mesenchymal stem cells (hMSCs) are a promising cell source for bone tissue engineering, but due to their limited number and donor variation, other cell types are used to answer relevant questions in bone tissue engineering. Since the extracellular matrix (ECM) is a complex entity with

  19. Extracellular matrix of the human aortic media: an ultrastructural histochemical and immunohistochemical study of the adult aortic media

    NARCIS (Netherlands)

    Dingemans, K. P.; Teeling, P.; Lagendijk, J. H.; Becker, A. E.

    2000-01-01

    Aortic distensability is the key to normal aortic function and relates to the lamellar unit in the media. However, the organization of the extracellular matrix components in these lamellar units, which are largely responsible for the distensability, is insufficiently known, especially in the human.

  20. Proliferation and extracellular matrix synthesis of smooth muscle cells cultured from human coronary atherosclerotic and restenotic lesions

    NARCIS (Netherlands)

    D.C. MacLeod (Donald); B.H. Strauss (Bradley); J. Escaned (Javier); V.A.W.M. Umans (Victor); R-J. van Suylen (Robert-Jan); A. Verkerk (Anton); P.J. de Feyter (Pim); P.W.J.C. Serruys (Patrick); M. de Jong (Marcel)

    1994-01-01

    textabstractOBJECTIVES. The purpose of this study was to examine the proliferative capacity and extracellular matrix synthesis of human coronary plaque cells in vitro. BACKGROUND. Common to both primary atherosclerosis and restenosis are vascular smooth muscle cell proliferation and production of

  1. Knockdown of asporin affects transforming growth factor-β1-induced matrix synthesis in human intervertebral annulus cells

    Directory of Open Access Journals (Sweden)

    Xu Jiang

    2016-10-01

    Conclusion: Our results have verified a functional feedback loop between TGF-β1 and asporin in human intervertebral annulus cells indicating that TGF-β1-induced annulus matrix biosynthesis can be significantly upregulated by knockdown of asporin. Therefore, asporin could be a potential new therapeutic target and inhibition of asporin could be adopted to enhance the anabolic effect of TGF-β1 in human intervertebral annulus cells in degenerative IVD diseases.

  2. Towards a quantitative understanding of oxygen tension and cell density evolution in fibrin hydrogels.

    Science.gov (United States)

    Demol, Jan; Lambrechts, Dennis; Geris, Liesbet; Schrooten, Jan; Van Oosterwyck, Hans

    2011-01-01

    The in vitro culture of hydrogel-based constructs above a critical size is accompanied by problems of unequal cell distribution when diffusion is the primary mode of oxygen transfer. In this study, an experimentally-informed mathematical model was developed to relate cell proliferation and death inside fibrin hydrogels to the local oxygen tension in a quantitative manner. The predictive capacity of the resulting model was tested by comparing its outcomes to the density, distribution and viability of human periosteum derived cells (hPDCs) that were cultured inside fibrin hydrogels in vitro. The model was able to reproduce important experimental findings, such as the formation of a multilayered cell sheet at the hydrogel periphery and the occurrence of a cell density gradient throughout the hydrogel. In addition, the model demonstrated that cell culture in fibrin hydrogels can lead to complete anoxia in the centre of the hydrogel for realistic values of oxygen diffusion and consumption. A sensitivity analysis also identified these two parameters, together with the proliferation parameters of the encapsulated cells, as the governing parameters for the occurrence of anoxia. In conclusion, this study indicates that mathematical models can help to better understand oxygen transport limitations and its influence on cell behaviour during the in vitro culture of cell-seeded hydrogels. Copyright © 2010 Elsevier Ltd. All rights reserved.

  3. Three-Dimensional In Vitro Skin and Skin Cancer Models Based on Human Fibroblast-Derived Matrix.

    Science.gov (United States)

    Berning, Manuel; Prätzel-Wunder, Silke; Bickenbach, Jackie R; Boukamp, Petra

    2015-09-01

    Three-dimensional in vitro skin and skin cancer models help to dissect epidermal-dermal and tumor-stroma interactions. In the model presented here, normal human dermal fibroblasts isolated from adult skin self-assembled into dermal equivalents with their specific fibroblast-derived matrix (fdmDE) over 4 weeks. The fdmDE represented a complex human extracellular matrix that was stabilized by its own heterogeneous collagen fiber meshwork, largely resembling a human dermal in vivo architecture. Complemented with normal human epidermal keratinocytes, the skin equivalent (fdmSE) thereof favored the establishment of a well-stratified and differentiated epidermis and importantly allowed epidermal regeneration in vitro for at least 24 weeks. Moreover, the fdmDE could be used to study the features of cutaneous skin cancer. Complementing fdmDE with HaCaT cells in different stages of malignancy or tumor-derived cutaneous squamous cell carcinoma cell lines, the resulting skin cancer equivalents (fdmSCEs) recapitulated the respective degree of tumorigenicity. In addition, the fdmSCE invasion phenotypes correlated with their individual degree of tissue organization, disturbance in basement membrane organization, and presence of matrix metalloproteinases. Together, fdmDE-based models are well suited for long-term regeneration of normal human epidermis and, as they recapitulate tumor-specific growth, differentiation, and invasion profiles of cutaneous skin cancer cells, also provide an excellent human in vitro skin cancer model.

  4. Expression and correlation of matrix metalloproteinase-7 and interleukin-15 in human osteoarthritis.

    Science.gov (United States)

    Tao, Yulei; Qiu, Xianxing; Xu, Changbo; Sun, Bo; Shi, Changxiu

    2015-01-01

    To investigate the expression and correlation of matrix metalloproteinase (MMP)-7 and interleukin (IL)-15 in human osteoarthritis (OA). From October 2013 to December 2014, 30 patients with OA were enrolled. In addition, anther 30 patients with simple meniscus injury were collected as a control group. There were no significant differences in age and gender between the two groups. Articular cartilage tissue was obtained from both OA patients and control group patients. Protein, mRNA, and serum expression levels of MMP-7 and IL-15 in the both two groups were determined by immunohistochemical (IHC), in situ hybridization, and enzyme-linked immunosorbent assay (ELISA) assay, respectively. Additionally, correlation between MMP-7 and IL-15 expression level in cartilage tissue and serum was assessed using Pearson correlation analysis. Protein, mRNA, and serum expression levels of MMP-7 and IL-15 in patients with OA were all significantly increased in OA patients compared with the control group. Besides, there were strong positive relationships between articular MMP-7 level and serum MMP-7 level (R(2) = 0.573, P = 0.018), between articular IL-15 level and serum IL-15 level (R(2) = 0.861, P = 0.023), and between serum IL-15 level and serum MMP-7 level (R(2) = 0.602, P = 0.012). These results suggest that MMP-7 and IL-15 might play important roles in the pathogenesis of OA, and IL-15 and MMP-7 has positive correlation in OA.

  5. Localized ridge defect augmentation using human pericardium membrane and demineralized bone matrix.

    Science.gov (United States)

    Vidyadharan, Arun Kumar; Ravindran, Anjana

    2014-01-01

    Patient wanted to restore her lost teeth with implants in the lower left first molar and second premolar region. Cone beam computerized tomography (CBCT) revealed inadequate bone width and height around future implant sites. The extraction socket of second premolar area revealed inadequate socket healing with sparse bone fill after 4 months of extraction. To evaluate the clinical feasibility of using a collagen physical resorbable barrier made of human pericardium (HP) to augment localized alveolar ridge defects for the subsequent placement of dental implants. Ridge augmentation was done in the compromised area using Puros® demineralized bone matrix (DBM) Putty with chips and an HP allograft membrane. Horizontal (width) and vertical hard tissue measurements with CBCT were recorded on the day of ridge augmentation surgery, 4 month and 7 months follow-up. Intra oral periapical taken 1 year after implant installation showed minimal crestal bone loss. Bone volume achieved through guided bone regeneration was a gain of 4.8 mm horizontally (width) and 6.8 mm vertically in the deficient ridge within a period of 7 months following the procedure. The results suggested that HP Allograft membrane may be a suitable component for augmentation of localized alveolar ridge defects in conjunction with DBM with bone chips.

  6. CD14+ monocytes promote the immunosuppressive effect of human umbilical cord matrix stem cells

    International Nuclear Information System (INIS)

    Wang, Ding; Chen, Ke; Du, Wei Ting; Han, Zhi-Bo; Ren, He; Chi, Ying

    2010-01-01

    Here, the effect of CD14 + monocytes on human umbilical cord matrix stem cell (hUC-MSC)-mediated immunosuppression was studied in vitro. hUC-MSCs exerted a potent inhibitory effect on the proliferation and interferon-γ (IFN-γ) secretion capacities of CD4 + and CD8 + T cells in response to anti-CD3/CD28 stimulation. Transwell co-culture system revealed that the suppressive effect was primarily mediated by soluble factors. Addition of prostaglandin synthesis inhibitors (indomethacin or NS-398) almost completely abrogated the immunosuppression activity of hUC-MSCs, identifying prostaglandin E 2 (PGE 2 ) as an important soluble mediator. CD14 + monocytes were found to be able to enhance significantly the immunosuppressive effect of hUC-MSCs in a dose-dependent fashion. Moreover, the inflammatory cytokine IL-1β, either exogenously added or produced by CD14 + monocytes in culture, could trigger expression of high levels of PGE 2 by hUC-MSCs, whereas inclusion of the IL-1 receptor antagonist (IL-1RA) in the culture down-regulated not only PGE 2 expression, but also reversed the promotional effect of CD14 + monocytes and partially restored CD4 + and CD8 + T cell proliferation and IFN-γ secretion. Our data demonstrate an important role of monocytes in the hUC-MSC-induced immunomodulation, which may have important implications in future efforts to explore the clinical potentials of hUC-MSCs.

  7. Two-dimensional electrophoretic analysis of nuclear matrix proteins in human colon adenocarcinoma.

    Science.gov (United States)

    Toumpanaki, A; Baltatzis, G E; Gaitanarou, E; Seretis, E; Toumpanakis, C; Aroni, K; Kittas, Christos; Voloudakis-Baltatzis, I E

    2009-01-01

    The aim of the present study was to observe possible qualitative and quantitative expression differences between nuclear matrix proteins (NMPs) of human colon adenocarcinoma and their mirror biopsies, using the technique of two-dimensional gel electrophoresis, in order to identify the existence of specific NMP fingerprints for colon cancer. Colon tissues were examined ultrastructurally and NMPs were isolated biochemically, by serial extraction of lipids, soluble proteins, DNA, RNA, and intermediate filaments and were separated according to their isoelectric point (pI) and their molecular weight (MW) by high-resolution two-dimensional electrophoresis (2D). By comparing the 2D electropherograms of colon cancer tissues and mirror biopsy tissues we observed qualitative and quantitative expression differences between their NMPs but also a differentiation of NMP composition between the stages of malignancy. Moreover, despite the similarities between mirror biopsy samples, a highlight percentage of exception was observed. Electrophoretic results provided in this study demonstrated that the examined NMPs could be further investigated as potential markers for detection of colorectal cancer in an early stage, for the assessment of the disease progression, as well as useful tools for individual therapy and for preventing a possible recurrence of cancer and metastasis.

  8. A new synthetic matrix metalloproteinase inhibitor reduces human mesenchymal stem cell adipogenesis.

    Directory of Open Access Journals (Sweden)

    Dale B Bosco

    Full Text Available Development of adipose tissue requires the differentiation of less specialized cells, such as human mesenchymal stem cells (hMSCs, into adipocytes. Since matrix metalloproteinases (MMPs play critical roles in the cell differentiation process, we conducted investigations to determine if a novel mercaptosulfonamide-based MMP inhibitor (MMPI, YHJ-7-52, could affect hMSC adipogenic differentiation and lipid accumulation. Enzyme inhibition assays, adipogenic differentiation experiments, and quantitative PCR methods were employed to characterize this inhibitor and determine its effect upon adipogenesis. YHJ-7-52 reduced lipid accumulation in differentiated cells by comparable amounts as a potent hydroxamate MMPI, GM6001. However, YHJ-7-82, a non-inhibitory structural analog of YHJ-7-52, in which the zinc-binding thiol group is replaced by a hydroxyl group, had no effect on adipogenesis. The two MMPIs (YHJ-7-52 and GM6001 were also as effective in reducing lipid accumulation in differentiated cells as T0070907, an antagonist of peroxisome-proliferator activated receptor gamma (PPAR-gamma, at a similar concentration. PPAR-gamma is a typical adipogenic marker and a key regulatory protein for the transition of preadiopocyte to adipocyte. Moreover, MMP inhibition was able to suppress lipid accumulation in cells co-treated with Troglitazone, a PPAR-gamma agonist. Our results indicate that MMP inhibitors may be used as molecular tools for adipogenesis and obesity treatment research.

  9. Distribution volumes of macromolecules in human ovarian and endometrial cancers--effects of extracellular matrix structure.

    Science.gov (United States)

    Haslene-Hox, Hanne; Oveland, Eystein; Woie, Kathrine; Salvesen, Helga B; Tenstad, Olav; Wiig, Helge

    2015-01-01

    Elements of the extracellular matrix (ECM), notably collagen and glucosaminoglycans, will restrict part of the space available for soluble macromolecules simply because the molecules cannot occupy the same space. This phenomenon may influence macromolecular drug uptake. To study the influence of steric and charge effects of the ECM on the distribution volumes of macromolecules in human healthy and malignant gynecologic tissues we used as probes 15 abundant plasma proteins quantified by high-resolution mass spectrometry. The available distribution volume (VA) of albumin was increased in ovarian carcinoma compared with healthy ovarian tissue. Furthermore, VA of plasma proteins between 40 and 190 kDa decreased with size for endometrial carcinoma and healthy ovarian tissue, but was independent of molecular weight for the ovarian carcinomas. An effect of charge on distribution volume was only found in healthy ovaries, which had lower hydration and high collagen content, indicating that a condensed interstitium increases the influence of negative charges. A number of earlier suggested biomarker candidates were detected in increased amounts in malignant tissue, e.g., stathmin and spindlin-1, showing that interstitial fluid, even when unfractionated, can be a valuable source for tissue-specific proteins. We demonstrate that the distribution of abundant plasma proteins in the interstitium can be elucidated by mass spectrometry methods and depends markedly on hydration and ECM structure. Our data can be used in modeling of drug uptake, and give indications on ECM components to be targeted to increase the uptake of macromolecular substances. Copyright © 2015 the American Physiological Society.

  10. Inhibition of matrix metalloproteinases expression in human dental pulp cells by all-trans retinoic acid

    Institute of Scientific and Technical Information of China (English)

    Jin Man Kim; Sang Wook Kang; Su-Mi Shin; Duck Su Kim; Kyong-Kyu Choi; Eun-Cheol Kim; Sun-Young Kim

    2014-01-01

    All-trans retinoic acid (ATRA) inhibits matrix metalloproteinase (MMP)-2 and MMP-9 in synovial fibroblasts, skin fibroblasts, bronchoalveolar lavage cells and cancer cells, but activates MMP-9 in neuroblast and leukemia cells. Very little is known regarding whether ATRA can activate or inhibit MMPs in human dental pulp cells (HDPCs). The purpose of this study was to determine the effects of ATRA on the production and secretion of MMP-2 and-9 in HDPCs. The productions and messenger RNA (mRNA) expressions of MMP-2 and-9 were accessed by gelatin zymography and real-time polymerase chain reaction (PCR), respectively. ATRA was found to decrease MMP-2 level in a dose-dependent manner. Significant reduction in MMP-2 mRNA expression was also observed in HDPCs treated with 25 mmol?L21 ATRA. However, HDPCs treated with ATRA had no effect on the pattern of MMP-9 produced or secreted in either cell extracts or conditioned medium fractions. Taken together, ATRA had an inhibitory effect on MMP-2 expression in HDPCs, which suggests that ATRA could be a candidate as a medicament which could control the inflammation of pulp tissue in vital pulp therapy and regenerative endodontics.

  11. In vitro evaluation of the interactions between human corneal endothelial cells and extracellular matrix proteins

    International Nuclear Information System (INIS)

    Choi, Jin San; Kim, Eun Young; Kim, Min Jeong; Giegengack, Matthew; Khan, Faraaz A; Soker, Shay; Khang, Gilson

    2013-01-01

    The corneal endothelium is the innermost cell layer of the cornea and rests on Descemet's membrane consisting of various extracellular matrix (ECM) proteins which can directly affect the cellular behaviors such as cell adhesion, proliferation, polarity, morphogenesis and function. The objective of this study was to investigate the interactions between the ECM environment and human corneal endothelial cells (HCECs), with the ultimate goal to improve cell proliferation and function in vitro. To evaluate the interaction of HCECs with ECM proteins, cells were seeded on ECM-coated tissue culture dishes, including collagen type I (COL I), collagen type IV (COL IV), fibronectin (FN), FNC coating mix (FNC) and laminin (LM). Cell adhesion and proliferation of HCECs on each substratum and expression of CEC markers were studied. The results showed that HCECs plated on the COL I, COL IV, FN and FNC-coated plates had enhanced cell adhesion initially; the number for COL I, COL IV, FN and FNC was significantly higher than the control (P < 0.05). In addition, cells grown on ECM protein-coated dishes showed more compact cellular morphology and CEC marker expression compared to cells seeded on uncoated dishes. Collectively, our results suggest that an adequate ECM protein combination can provide a long-term culture environment for HCECs for corneal endothelium transplantation. (paper)

  12. Structure and self-assembly of the calcium binding matrix protein of human metapneumovirus.

    Science.gov (United States)

    Leyrat, Cedric; Renner, Max; Harlos, Karl; Huiskonen, Juha T; Grimes, Jonathan M

    2014-01-07

    The matrix protein (M) of paramyxoviruses plays a key role in determining virion morphology by directing viral assembly and budding. Here, we report the crystal structure of the human metapneumovirus M at 2.8 Å resolution in its native dimeric state. The structure reveals the presence of a high-affinity Ca²⁺ binding site. Molecular dynamics simulations (MDS) predict a secondary lower-affinity site that correlates well with data from fluorescence-based thermal shift assays. By combining small-angle X-ray scattering with MDS and ensemble analysis, we captured the structure and dynamics of M in solution. Our analysis reveals a large positively charged patch on the protein surface that is involved in membrane interaction. Structural analysis of DOPC-induced polymerization of M into helical filaments using electron microscopy leads to a model of M self-assembly. The conservation of the Ca²⁺ binding sites suggests a role for calcium in the replication and morphogenesis of pneumoviruses. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  13. Enhanced expression of two discrete isoforms of matrix metalloproteinase-2 in experimental and human diabetic nephropathy.

    Directory of Open Access Journals (Sweden)

    Sang Soo Kim

    Full Text Available We recently reported on the enhanced expression of two isoforms of matrix metalloproteinase-2 (MMP-2 in human renal transplantation delayed graft function. These consist of the conventional secreted, full length MMP-2 isoform (FL-MMP-2 and a novel intracellular N-Terminal Truncated isoform (NTT-MMP-2 generated by oxidative stress-mediated activation of an alternate promoter in the MMP-2 first intron. Here we evaluated the effect of hyperglycemia and diabetes mellitus on the in vitro and in vivo expression of the two MMP-2 isoforms.We quantified the abundance of the FL-MMP-2 and NTT-MMP-2 transcripts by qPCR in HK2 cells cultured in high glucose or 4-hydroxy-2-hexenal (HHE and tested the effects of the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC. The streptozotocin (STZ murine model of Type I diabetes mellitus and renal biopsies of human diabetic nephropathy were used in this study.Both isoforms of MMP-2 in HK2 cells were upregulated by culture in high glucose or with HHE. PDTC treatment did not suppress high glucose-mediated FL-MMP-2 expression but potently inhibited NTT-MMP-2 expression. With STZ-treated mice, renal cortical expression of both isoforms was increased (FL-MMP-2, 1.8-fold; NTT-MMP-2, greater than 7-fold. Isoform-specific immunohistochemical staining revealed low, but detectable levels of the FL-MMP-2 isoform in controls, while NTT-MMP-2 was not detected. While there was a modest increase in tubular epithelial cell staining for FL-MMP-2 in STZ-treated mice, NTT-MMP-2 was intensely expressed in a basolateral pattern. FL-MMP-2 and NTT-MMP-2 isoform expression as quantified by qPCR were both significantly elevated in renal biopsies of human diabetic nephropathy (12-fold and 3-fold, respectively.The expression of both isoforms of MMP-2 was enhanced in an experimental model of diabetic nephropathy and in human diabetic nephropathy. Selective MMP-2 isoform inhibition could offer a novel approach for the treatment of diabetic renal

  14. Immunosuppressive function of mesenchymal stem cells from human umbilical cord matrix in immune thrombocytopenia patients.

    Science.gov (United States)

    Ma, Li; Zhou, Zeping; Zhang, Donglei; Yang, Shaoguang; Wang, Jinhong; Xue, Feng; Yang, Yanhui; Yang, Renchi

    2012-05-01

    Human umbilical cord matrix/Wharton's jelly (hUC)-derived mesenchymal stem cells (MSC) have been shown to have marked therapeutic effects in a number of inflammatory diseases and autoimmune diseases in humans based on their potential for immunosuppression and their low immunogenicity. Currently, no data are available on the effectiveness of UC-MSC transplantation in immune thrombocytopenia (ITP) patients. It was the objective of this study to assess the effect of allogeneic UC-MSCs on ITP patients in vitro and in vivo. Peripheral blood mononuclear cells (PBMCs) and bone marrow mononuclear cells (BM-MNCs) from ITP patients and healthy controls were co-cultured with UC-MSCs for three days and seven days, respectively. Flow cytometry and ELISA were applied to assess the various parameters. In PBMCs from ITP patients, the proliferation of autoreactive T, B lymphocytes and destruction of autologous platelets were dramatically suppressed by UC-MSCs. UC-MSCs not only suppressed co-stimulatory molecules CD80, CD40L and FasL expression but also in shifting Th1/Th2/Treg cytokines profile in ITP patients. UC-MSCs obviously reversed the dysfunctions of megakaryocytes by promoting platelet production and decreasing the number of living megakaryocytes as well as early apoptosis. In addition, the level of thrombopoietin was increased significantly. Our clinical study showed that UC-MSCs play a role in alleviating refractory ITP by increasing platelet numbers. These findings suggested that UC-MSCs transplantation might be a potential therapy for ITP.

  15. Bone induction by composites of bioresorbable carriers and demineralized bone in rats: a comparative study of fibrin-collagen paste, fibrin sealant, and polyorthoester with gentamicin

    DEFF Research Database (Denmark)

    Pinholt, E M; Solheim, E; Bang, G

    1992-01-01

    fibrin-collagen paste and fibrin sealant inhibited bone induction and produced a chronic inflammation; part of the fibrin-collagen paste was still present at 4 weeks. Polyorthoester with gentamicin was almost completely absorbed, induced minimal tissue reaction, and did not inhibit osteoinduction....

  16. Expression of extracellular matrix components and related growth factors in human tendon and muscle after acute exercise

    DEFF Research Database (Denmark)

    Heinemeier, K M; Bjerrum, S S; Schjerling, P

    2013-01-01

    Acute kicking exercise induces collagen synthesis in both tendon and muscle in humans, but it is not known if this relates to increased collagen transcription and if other matrix genes are regulated. Young men performed 1 h of one-leg kicking at 67% of max workload. Biopsies were taken from...... the patellar tendon and vastus lateralis muscle of each leg at 2 (n = 10), 6 (n = 11), or 26 h (n = 10) after exercise. Levels of messenger ribonucleic acid mRNA for collagens, noncollagenous matrix proteins, and growth factors were measured with real-time reverse transcription polymerase chain reaction...

  17. Extracellular matrix metabolism disorder induced by mechanical strain on human parametrial ligament fibroblasts.

    Science.gov (United States)

    Min, Jie; Li, Bingshu; Liu, Cheng; Guo, Wenjun; Hong, Shasha; Tang, Jianming; Hong, Li

    2017-05-01

    Pelvic organ prolapse (POP) is a global health problem that may seriously impact the quality of life of the sufferer. The present study aimed to investigate the potential mechanisms underlying alterations in extracellular matrix (ECM) metabolism in the pathogenesis of POP, by investigating the expression of ECM components in human parametrial ligament fibroblasts (hPLFs) subject to various mechanical strain loads. Fibroblasts derived from parametrial ligaments were cultured from patients with POP and without malignant tumors, who underwent vaginal hysterectomy surgery. Fibroblasts at generations 3‑6 of exponential phase cells were selected, and a four‑point bending device was used for 0, 1,333 or 5,333 µ mechanical loading of cells at 0.5 Hz for 4 h. mRNA and protein expression levels of collagen type I α 1 chain (COL1A1), collagen type III α 1 chain (COL3A1), elastin, matrix metalloproteinase (MMP) ‑2 and ‑9, and transforming growth factor (TGF)‑β1 were detected by reverse transcription‑quantitative polymerase chain reaction and western blotting, respectively. Under increased mechanical strain (5,333 µ), mRNA and protein expression levels of COL1A1, COL3A1 elastin and TGF‑β1 decreased, particularly COL1A1; however, mRNA and protein expression levels of MMP‑2 and ‑9 were significantly increased, compared with the control group (0 µ strain). Following 1,333 µ mechanical strain, mRNA and protein expression levels of COL1A1, COL3A1 elastin and MMP‑2 increased, and MMP‑9 decreased, whereas no significant differences were observed in TGF‑β1 mRNA and protein expression levels. In conclusion, ECM alterations may be involved in pathogenesis of POP, with decreased synthesis and increased degradation of collagen and elastin. Furthermore, the TGF‑β1 signaling pathway may serve an important role in this process and thus may supply a new target and strategy for understanding the etiology and therapy of POP.

  18. Microneural anastomosis with fibrin glue : an experimental study.

    OpenAIRE

    Suri A; Mehta V; Sarkar C

    2002-01-01

    An experimental study was designed to compare the histological analysis of nerve anastomosis with 10-0 microsurgical sutures and fibrin adhesive. Wistar albino rats′ sciatic nerves were transected and repaired either with fibrin adhesive-Beriplast P (M/s Centeon-Cadila Health Care) or with 10-0 monofilament microsutures. Histological assessment was performed at 10, 20, 30, 60 and 90 days after surgery. Functional recovery of the sciatic nerves started at two months and was near normal ...

  19. Submillisecond elastic recoil reveals molecular origins of fibrin fiber mechanics.

    Science.gov (United States)

    Hudson, Nathan E; Ding, Feng; Bucay, Igal; O'Brien, E Timothy; Gorkun, Oleg V; Superfine, Richard; Lord, Susan T; Dokholyan, Nikolay V; Falvo, Michael R

    2013-06-18

    Fibrin fibers form the structural scaffold of blood clots. Thus, their mechanical properties are of central importance to understanding hemostasis and thrombotic disease. Recent studies have revealed that fibrin fibers are elastomeric despite their high degree of molecular ordering. These results have inspired a variety of molecular models for fibrin's elasticity, ranging from reversible protein unfolding to rubber-like elasticity. An important property that has not been explored is the timescale of elastic recoil, a parameter that is critical for fibrin's mechanical function and places a temporal constraint on molecular models of fiber elasticity. Using high-frame-rate imaging and atomic force microscopy-based nanomanipulation, we measured the recoil dynamics of individual fibrin fibers and found that the recoil was orders of magnitude faster than anticipated from models involving protein refolding. We also performed steered discrete molecular-dynamics simulations to investigate the molecular origins of the observed recoil. Our results point to the unstructured αC regions of the otherwise structured fibrin molecule as being responsible for the elastic recoil of the fibers. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  20. Scaffold-assisted cartilage tissue engineering using infant chondrocytes from human hip cartilage.

    Science.gov (United States)

    Kreuz, P C; Gentili, C; Samans, B; Martinelli, D; Krüger, J P; Mittelmeier, W; Endres, M; Cancedda, R; Kaps, C

    2013-12-01

    Studies about cartilage repair in the hip and infant chondrocytes are rare. The aim of our study was to evaluate the use of infant articular hip chondrocytes for tissue engineering of scaffold-assisted cartilage grafts. Hip cartilage was obtained from five human donors (age 1-10 years). Expanded chondrocytes were cultured in polyglycolic acid (PGA)-fibrin scaffolds. De- and re-differentiation of chondrocytes were assessed by histological staining and gene expression analysis of typical chondrocytic marker genes. In vivo, cartilage matrix formation was assessed by histology after subcutaneous transplantation of chondrocyte-seeded PGA-fibrin scaffolds in immunocompromised mice. The donor tissue was heterogenous showing differentiated articular cartilage and non-differentiated tissue and considerable expression of type I and II collagens. Gene expression analysis showed repression of typical chondrocyte and/or mesenchymal marker genes during cell expansion, while markers were re-induced when expanded cells were cultured in PGA-fibrin scaffolds. Cartilage formation after subcutaneous transplantation of chondrocyte loaded PGA-fibrin scaffolds in nude mice was variable, with grafts showing resorption and host cell infiltration or formation of hyaline cartilage rich in type II collagen. Addition of human platelet rich plasma (PRP) to cartilage grafts resulted robustly in formation of hyaline-like cartilage that showed type II collagen and regions with type X collagen. These results suggest that culture of expanded and/or de-differentiated infant hip cartilage cells in PGA-fibrin scaffolds initiates chondrocyte re-differentiation. The heterogenous donor tissue containing immature chondrocytes bears the risk of cartilage repair failure in vivo, which may be possibly overcome by the addition of PRP. Copyright © 2013 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

  1. Release of tensile strain on engineered human tendon tissue disturbs cell adhesions, changes matrix architecture, and induces an inflammatory phenotype

    DEFF Research Database (Denmark)

    Bayer, Monika L; Schjerling, Peter; Herchenhan, Andreas

    2014-01-01

    Mechanical loading of tendon cells results in an upregulation of mechanotransduction signaling pathways, cell-matrix adhesion and collagen synthesis, but whether unloading removes these responses is unclear. We investigated the response to tension release, with regard to matrix proteins, pro......-inflammatory mediators and tendon phenotypic specific molecules, in an in vitro model where tendon-like tissue was engineered from human tendon cells. Tissue sampling was performed 1, 2, 4 and 6 days after surgical de-tensioning of the tendon construct. When tensile stimulus was removed, integrin type collagen receptors...... were upregulated. Stimulation with the cytokine TGF-β1 had distinct effects on some tendon-related genes in both tensioned and de-tensioned tissue. These findings indicate an important role of mechanical loading for cellular and matrix responses in tendon, including that loss of tension leads...

  2. Sequential changes in histologic pattern and extracellular matrix deposition during the healing of chronic venous ulcers.

    OpenAIRE

    Herrick, S. E.; Sloan, P.; McGurk, M.; Freak, L.; McCollum, C. N.; Ferguson, M. W.

    1992-01-01

    As part of a major clinical trial, sequential biopsies were taken from the margins of venous leg ulcers during their healing. The changing patterns of tissue architecture and extracellular matrix synthesis during healing were documented histologically and immunocytochemically. Initial biopsies were similar in appearance: prominent fibrin cuffs, variable inflammation, hemosiderin, and red blood cell extravasation. So called "fibrin cuffs" were highly organized structures composed of laminin, f...

  3. A CCR2 macrophage endocytic pathway mediates extravascular fibrin clearance in vivo

    DEFF Research Database (Denmark)

    Motley, Michael P; Madsen, Daniel H; Jürgensen, Henrik J

    2016-01-01

    cellular endocytosis and lysosomal targeting, revealing a novel intracellular pathway for extravascular fibrin degradation. A C-C chemokine receptor type 2 (CCR2)-positive macrophage subpopulation constituted the majority of fibrin-uptaking cells. Consequently, cellular fibrin uptake was diminished...... by elimination of CCR2-expressing cells. The CCR2-positive macrophage subtype was different from collagen-internalizing M2-like macrophages. Cellular fibrin uptake was strictly dependent on plasminogen and plasminogen activator. Surprisingly, however, fibrin endocytosis was unimpeded by the absence of the fibrin...... subsets of macrophages employing distinct molecular pathways....

  4. Oxidation modifies the structure and function of the extracellular matrix generated by human coronary artery endothelial cells.

    Science.gov (United States)

    Chuang, Christine Y; Degendorfer, Georg; Hammer, Astrid; Whitelock, John M; Malle, Ernst; Davies, Michael J

    2014-04-15

    ECM (extracellular matrix) materials, such as laminin, perlecan, type IV collagen and fibronectin, play a key role in determining the structure of the arterial wall and the properties of cells that interact with the ECM. The aim of the present study was to investigate the effect of peroxynitrous acid, an oxidant generated by activated macrophages, on the structure and function of the ECM laid down by HCAECs (human coronary artery endothelial cells) in vitro and in vivo. We show that exposure of HCAEC-derived native matrix components to peroxynitrous acid (but not decomposed oxidant) at concentrations >1 μM results in a loss of antibody recognition of perlecan, collagen IV, and cell-binding sites on laminin and fibronectin. Loss of recognition was accompanied by decreased HCAEC adhesion. Real-time PCR showed up-regulation of inflammation-associated genes, including MMP7 (matrix metalloproteinase 7) and MMP13, as well as down-regulation of the laminin α2 chain, in HCAECs cultured on peroxynitrous acid-treated matrix compared with native matrix. Immunohistochemical studies provided evidence of co-localization of laminin with 3-nitrotyrosine, a biomarker of peroxynitrous acid damage, in type II-III/IV human atherosclerotic lesions, consistent with matrix damage occurring during disease development in vivo. The results of the present study suggest a mechanism through which peroxynitrous acid modifies endothelial cell-derived native ECM proteins of the arterial basement membrane in atherosclerotic lesions. These changes to ECM and particularly perlecan and laminin may be important in inducing cellular dysfunction and contribute to atherogenesis.

  5. Heparan sulfate proteoglycan from the extracellular matrix of human lung fibroblasts. Isolation, purification, and core protein characterization

    International Nuclear Information System (INIS)

    Heremans, A.; Cassiman, J.J.; Van den Berghe, H.; David, G.

    1988-01-01

    Confluent cultured human lung fibroblasts were labeled with 35SO4(2-). After 48 h of labeling, the pericellular matrix was prepared by Triton X-100 and deoxycholate extraction of the monolayers. Heparan sulfate proteoglycan (HSPG) accounted for nearly 80% of the total matrix [35S]proteoglycans. After solubilization in 6 M guanidinium HCl and cesium chloride density gradient centrifugation, the majority (78%) of these [35S] HSPG equilibrated at an average buoyant density of 1.35 g/ml. This major HSPG fraction was purified by ion-exchange chromatography on Mono Q and by gel filtration on Sepharose CL-4B, and further characterized by gel electrophoresis and immunoblotting. Intact [35S]HSPG eluted with Kav 0.1 from Sepharose CL-4B, whereas the protein-free [35S]heparan sulfate chains, obtained by alkaline borohydride treatment of the proteoglycan fractions, eluted with Kav 0.45 (Mr approximately 72,000). When analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography, core (protein) preparations, obtained by heparitinase digestion of 125I-labeled HSPG fractions, yielded one major labeled band with apparent molecular mass of approximately 300 kDa. Reduction with beta-mercaptoethanol slightly increased the apparent Mr of the labeled band, suggesting a single polypeptide structure and the presence of intrachain disulfide bonds. Immunoadsorption experiments and immunostaining of electrophoretically separated heparitinase-digested core proteins with monoclonal antibodies raised against matrix and cell surface-associated HSPG suggested that the major matrix-associated HSPG of cultured human lung fibroblasts is distinct from the HSPG that are anchored in the membranes of these cells. Binding studies suggested that this matrix HSPG interacts with several matrix components, both through its glycosaminoglycan chains and through its heparitinase-resistant core. (Abstract Truncated)

  6. Matrix Degradation in Human Immunodeficiency Virus Type 1-Associated Tuberculosis and Tuberculosis Immune Reconstitution Inflammatory Syndrome: A Prospective Observational Study.

    Science.gov (United States)

    Walker, Naomi F; Wilkinson, Katalin A; Meintjes, Graeme; Tezera, Liku B; Goliath, Rene; Peyper, Janique M; Tadokera, Rebecca; Opondo, Charles; Coussens, Anna K; Wilkinson, Robert J; Friedland, Jon S; Elkington, Paul T

    2017-07-01

    Extensive immunopathology occurs in human immunodeficiency virus (HIV)/tuberculosis (TB) coinfection, but the underlying molecular mechanisms are not well-defined. Excessive matrix metalloproteinase (MMP) activity is emerging as a key process but has not been systematically studied in HIV-associated TB. We performed a cross-sectional study of matrix turnover in HIV type 1 (HIV-1)-infected and -uninfected TB patients and controls, and a prospective cohort study of HIV-1-infected TB patients at risk of TB immune reconstitution inflammatory syndrome (TB-IRIS), in Cape Town, South Africa. Sputum and plasma MMP concentrations were quantified by Luminex, plasma procollagen III N-terminal propeptide (PIIINP) by enzyme-linked immunosorbent assay, and urinary lipoarabinomannan (LAM) by Alere Determine TB LAM assay. Peripheral blood mononuclear cells from healthy donors were cultured with Mycobacterium tuberculosis and extracellular matrix in a 3D model of TB granuloma formation. MMP activity differed between HIV-1-infected and -uninfected TB patients and corresponded with specific TB clinical phenotypes. HIV-1-infected TB patients had reduced pulmonary MMP concentrations, associated with reduced cavitation, but increased plasma PIIINP, compared to HIV-1-uninfected TB patients. Elevated extrapulmonary extracellular matrix turnover was associated with TB-IRIS, both before and during TB-IRIS onset. The predominant collagenase was MMP-8, which was likely neutrophil derived and M. tuberculosis-antigen driven. Mycobacterium tuberculosis-induced matrix degradation was suppressed by the MMP inhibitor doxycycline in vitro. MMP activity in TB differs by HIV-1 status and compartment, and releases matrix degradation products. Matrix turnover in HIV-1-infected patients is increased before and during TB-IRIS, informing novel diagnostic strategies. MMP inhibition is a potential host-directed therapy strategy for prevention and treatment of TB-IRIS. © The Author 2017. Published by Oxford

  7. Matrix metalloproteinase-2 of human carotid atherosclerotic plaques promotes platelet activation. Correlation with ischaemic events.

    Science.gov (United States)

    Lenti, Massimo; Falcinelli, Emanuela; Pompili, Marcella; de Rango, Paola; Conti, Valentina; Guglielmini, Giuseppe; Momi, Stefania; Corazzi, Teresa; Giordano, Giuseppe; Gresele, Paolo

    2014-06-01

    Purified active matrix metalloproteinase-2 (MMP-2) is able to promote platelet aggregation. We aimed to assess the role of MMP-2 expressed in atherosclerotic plaques in the platelet-activating potential of human carotid plaques and its correlation with ischaemic events. Carotid plaques from 81 patients undergoing endarterectomy were tested for pro-MMP-2 and TIMP-2 content by zymography and ELISA. Plaque extracts were incubated with gel-filtered platelets from healthy volunteers for 2 minutes before the addition of a subthreshold concentration of thrombin receptor activating peptide-6 (TRAP-6) and aggregation was assessed. Moreover, platelet deposition on plaque extracts immobilised on plastic coverslips under high shear-rate flow conditions was measured. Forty-three plaque extracts (53%) potentiated platelet aggregation (+233 ± 26.8%), an effect prevented by three different specific MMP-2 inhibitors (inhibitor II, TIMP-2, moAb anti-MMP-2). The pro-MMP-2/TIMP-2 ratio of plaques potentiating platelet aggregation was significantly higher than that of plaques not potentiating it (3.67 ± 1.21 vs 1.01 ± 0.43, p<0.05). Moreover, the platelet aggregation-potentiating effect, the active-MMP-2 content and the active MMP-2/pro-MMP-2 ratio of plaque extracts were significantly higher in plaques from patients who developed a subsequent major cardiovascular event. In conclusion, atherosclerotic plaques exert a prothrombotic effect by potentiating platelet activation due to their content of MMP-2; an elevated MMP-2 activity in plaques is associated with a higher rate of subsequent ischaemic cerebrovascular events.

  8. Outcomes of arthroscopic revision rotator cuff repair with acellular human dermal matrix allograft augmentation.

    Science.gov (United States)

    Hohn, Eric A; Gillette, Blake P; Burns, Joseph P

    2018-05-01

    The purpose was to assess the minimum 2-year patient-reported outcomes and failure rate of patients who underwent revision arthroscopic rotator cuff repair augmented with acellular human dermal matrix (AHDM) allograft for repairable retears. From 2008-2014, patients who underwent revision rotator cuff repair augmented with AHDM with greater than 2 years' follow-up by a single surgeon were retrospectively reviewed. Data regarding surgical history, demographic characteristics, and medical comorbidities were collected. Outcome data included American Shoulder and Elbow Surgeons (ASES) and Single Assessment Numeric Evaluation (SANE) scores, as well as rotator cuff healing on magnetic resonance imaging or ultrasound. Retears and subsequent surgical procedures were characterized. A total of 28 patients met our inclusion criteria, and 23 (82%) were available for follow-up at 2 years. The mean age was 60.1 ± 9.3 years (range, 43-79 years), with a mean follow-up period of 48 ± 23 months. All patients had at least 1 prior rotator cuff repair. Of the 23 patients, 13 (56%) underwent postoperative imaging, and 4 of these 13 (31%) had a retear. A reoperation was performed in 3 of 23 patients (13%). Among the 6 patients with both preoperative and postoperative outcome scores, we saw improvement in the ASES score from 56 to 85 (P = .03) and in the SANE score from 42 to 76 (P = .03). The full cohort's mean postoperative ASES and SANE scores were 77 and 69, respectively. AHDM allograft augmentation is a safe and effective treatment method for patients with full-thickness rotator cuff retears. Further research is needed with larger studies to confirm these findings from our small cohort of patients. Copyright © 2017 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.

  9. Regulation of human mesenchymal stem cells differentiation into chondrocytes in extracellular matrix-based hydrogel scaffolds.

    Science.gov (United States)

    Du, Mingchun; Liang, Hui; Mou, Chenchen; Li, Xiaoran; Sun, Jie; Zhuang, Yan; Xiao, Zhifeng; Chen, Bing; Dai, Jianwu

    2014-02-01

    To induce human mesenchymal stem cells (hMSCs) to differentiate into chondrocytes in three-dimensional (3D) microenvironments, we developed porous hydrogel scaffolds using the cartilage extracellular matrix (ECM) components of chondroitin sulfate (CS) and collagen (COL). The turbidity and viscosity experiments indicated hydrogel could form through pH-triggered co-precipitation when pH=2-3. Enzyme-linked immunosorbent assay (ELISA) confirmed the hydrogel scaffolds could controllably release growth factors as envisaged. Transforming growth factor-β (TGF-β) was released to stimulate hMSCs differentiation into chondrocytes; and then collagen binding domain-basic fibroblast growth factor (CBD-bFGF) was released to improve the differentiation and preserve the chondrocyte phenotype. In in vitro cell culture experiments, the differentiation processes were compared in different microenvironments: 2D culture in culture plate as control, 3D culture in the fabricated scaffolds without growth factors (CC), the samples with CBD-bFGF (CC-C), the samples with TGF-β (CC-T), the samples with CBD-bFGF/TGF-β (CC-CT). Real-time polymerase chain reaction (RT-PCR) revealed the hMSC marker genes of CD44 and CD105 decreased; at the same time the chondrocyte marker genes of collagen type II and aggrecan increased, especially in the CC-CT sample. Immunostaining results further confirmed the hMSC marker protein of CD 44 disappeared and the chondrocyte marker protein of collagen type II emerged over time in the CC-CT sample. These results imply the ECM-based hydrogel scaffolds with growth factors can supply suitable 3D cell niches for hMSCs differentiation into chondrocytes and the differentiation process can be regulated by the controllably released growth factors. Copyright © 2013 Elsevier B.V. All rights reserved.

  10. Levels of matrix metalloproteinase-7 and osteopontin in human gingival crevicular fluid during initial tooth movement

    Directory of Open Access Journals (Sweden)

    Dhaval Oswal

    2015-01-01

    Full Text Available Purpose: During orthodontic treatment, the early response of periodontal tissues to mechanical stress involves several metabolic changes that allow tooth movement. The purpose of this investigation was to evaluate osteopontin (OPN and matrix metalloproteinase (MMP-7 in the gingival crevicular fluid (GCF of human teeth exposed to orthodontic force. Materials and Methods: GCF samples were obtained from 15 healthy orthodontic patients (age, 12-22 years. In each patient, the left maxillary canine having the fixed orthodontic appliance was used as the test tooth, and its antagonist, with no appliance, was the control. Orthodontic force, 75 g was applied using a 16 × 22 beta titanium closing loop. The GCF sampling on the disto-buccal aspects of experimental and control tooth was performed at specific time interval with sterilized absorbent paper point. Processing was carried out with enzyme-linked immunosorbent assay to detect OPN and MMP-7 levels. Results: The peak level of OPN was seen after 1 h application of orthodontic force which was 1280.36 pg/ml ± 185.02. The peak level of MMP-7 was seen at 0 h which was 598.3 pg/ml ± 107.5. The levels of OPN after 1 h increased to 1280.36 pg/ml ± 185.02, and they decreased at 24 h to 1012.86 pg/ml ± 168.47 (P = 0.001. The levels of MMP-7 after 1 h decreased to 478 pg/ml ± 99.7 which increased at 24 h to 526.9 pg/ml ± 99.2. Conclusions: Orthodontic forces affect both OPN and MMP-7 protein levels on the compression side in a time-dependent fashion.

  11. CD14{sup +} monocytes promote the immunosuppressive effect of human umbilical cord matrix stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Ding, E-mail: qqhewd@gmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Chen, Ke, E-mail: chenke_59@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Du, Wei Ting, E-mail: duwtpumc@yahoo.com.cn [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); Han, Zhi-Bo, E-mail: zhibohan@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Ren, He, E-mail: knifesharp2000@hotmail.com [National Engineering Research Center of Cell Products, AmCellGene Co. Ltd, TEDA, Tianjin (China); Chi, Ying, E-mail: caizhuying@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); and others

    2010-09-10

    Here, the effect of CD14{sup +} monocytes on human umbilical cord matrix stem cell (hUC-MSC)-mediated immunosuppression was studied in vitro. hUC-MSCs exerted a potent inhibitory effect on the proliferation and interferon-{gamma} (IFN-{gamma}) secretion capacities of CD4{sup +} and CD8{sup +} T cells in response to anti-CD3/CD28 stimulation. Transwell co-culture system revealed that the suppressive effect was primarily mediated by soluble factors. Addition of prostaglandin synthesis inhibitors (indomethacin or NS-398) almost completely abrogated the immunosuppression activity of hUC-MSCs, identifying prostaglandin E{sub 2} (PGE{sub 2}) as an important soluble mediator. CD14{sup +} monocytes were found to be able to enhance significantly the immunosuppressive effect of hUC-MSCs in a dose-dependent fashion. Moreover, the inflammatory cytokine IL-1{beta}, either exogenously added or produced by CD14{sup +} monocytes in culture, could trigger expression of high levels of PGE{sub 2} by hUC-MSCs, whereas inclusion of the IL-1 receptor antagonist (IL-1RA) in the culture down-regulated not only PGE{sub 2} expression, but also reversed the promotional effect of CD14{sup +} monocytes and partially restored CD4{sup +} and CD8{sup +} T cell proliferation and IFN-{gamma} secretion. Our data demonstrate an important role of monocytes in the hUC-MSC-induced immunomodulation, which may have important implications in future efforts to explore the clinical potentials of hUC-MSCs.

  12. Human disc cells in monolayer vs 3D culture: cell shape, division and matrix formation

    Directory of Open Access Journals (Sweden)

    Hanley Edward N

    2000-10-01

    Full Text Available Abstract Background The relationship between cell shape, proliferation, and extracellular matrix (ECM production, important aspects of cell behavior, is examined in a little-studied cell type, the human annulus cell from the intervertebral disc, during monolayer vs three-dimensional (3D culture. Results Three experimental studies showed that cells respond specifically to culture microenvironments by changes in cell shape, mitosis and ECM production: 1 Cell passages showed extensive immunohistochemical evidence of Type I and II collagens only in 3D culture. Chondroitin sulfate and keratan sulfate were abundant in both monolayer and 3D cultures. 2 Cells showed significantly greater proliferation in monolayer in the presence of platelet-derived growth factor compared to cells in 3D. 3 Cells on Matrigel™-coated monolayer substrates became rounded and formed nodular colonies, a finding absent during monolayer growth. Conclusions The cell's in vivo interactions with the ECM can regulate shape, gene expression and other cell functions. The shape of the annulus cell changes markedly during life: the young, healthy disc contains spindle shaped cells and abundant collagen. With aging and degeneration, many cells assume a strikingly different appearance, become rounded and are surrounded by unusual accumulations of ECM products. In vitro manipulation of disc cells provides an experimental window for testing how disc cells from given individuals respond when they are grown in environments which direct cells to have either spindle- or rounded-shapes. In vitro assessment of the response of such cells to platelet-derived growth factor and to Matrigel™ showed a continued influence of cell shape even in the presence of a growth factor stimulus. These findings contribute new information to the important issue of the influence of cell shape on cell behavior.

  13. Matrix metalloproteinase-12 gene regulation by a PPAR alpha agonist in human monocyte-derived macrophages

    International Nuclear Information System (INIS)

    Souissi, Imen Jguirim; Billiet, Ludivine; Cuaz-Perolin, Clarisse; Slimane, Mohamed-Naceur; Rouis, Mustapha

    2008-01-01

    MMP-12, a macrophage-specific matrix metalloproteinase with large substrate specificity, has been reported to be highly expressed in mice, rabbits and human atherosclerotic lesions. Increased MMP-12 from inflammatory macrophages is associated with several degenerative diseases such as atherosclerosis. In this manuscript, we show that IL-1β, a proinflammatory cytokine found in atherosclerotic plaques, increases both mRNA and protein levels of MMP-12 in human monocyte-derived macrophages (HMDM). Since peroxisome proliferator-activated receptors (PPARs), such as PPARα and PPARγ, are expressed in macrophages and because PPAR activation exerts an anti-inflammatory effect on vascular cells, we have investigated the effect of PPARα and γ isoforms on MMP-12 regulation in HMDM. Our results show that MMP-12 expression (mRNA and protein) is down regulated in IL-1β-treated macrophages only in the presence of a specific PPARα agonist, GW647, in a dose-dependent manner. In contrast, this inhibitory effect was abolished in IL-1β-stimulated peritoneal macrophages isolated from PPARα -/- mice and treated with the PPARα agonist, GW647. Moreover, reporter gene transfection experiments using different MMP-12 promoter constructs showed a reduction of the promoter activities by ∼ 50% in IL-1β-stimulated PPARα-pre-treated cells. However, MMP-12 promoter analysis did not reveal the presence of a PPRE response element. The IL-1β effect is known to be mediated through the AP-1 binding site. Mutation of the AP-1 site, located at - 81 in the MMP-12 promoter region relative to the transcription start site, followed by transfection analysis, gel shift and ChIP experiments revealed that the inhibitory effect was the consequence of the protein-protein interaction between GW 647-activated PPARα and c-Fos or c-Jun transcription factors, leading to inhibition of their binding to the AP-1 motif. These studies suggest that PPARα agonists may be used therapeutically, not only for lipid

  14. Fibrin Gels Exhibit Improved Biological, Structural, and Mechanical Properties Compared with Collagen Gels in Cell-Based Tendon Tissue-Engineered Constructs

    Science.gov (United States)

    Dyment, Nathaniel A.; Lu, Yinhui; Rao, Marepalli; Shearn, Jason T.; Rowe, David W.; Kadler, Karl E.; Butler, David L.

    2015-01-01

    The prevalence of tendon and ligament injuries and inadequacies of current treatments is driving the need for alternative strategies such as tissue engineering. Fibrin and collagen biopolymers have been popular materials for creating tissue-engineered constructs (TECs), as they exhibit advantages of biocompatibility and flexibility in construct design. Unfortunately, a few studies have directly compared these materials for tendon and ligament applications. Therefore, this study aims at determining how collagen versus fibrin hydrogels affect the biological, structural, and mechanical properties of TECs during formation in vitro. Our findings show that tendon and ligament progenitor cells seeded in fibrin constructs exhibit improved tenogenic gene expression patterns compared with their collagen-based counterparts for approximately 14 days in culture. Fibrin-based constructs also exhibit improved cell-derived collagen alignment, increased linear modulus (2.2-fold greater) compared with collagen-based constructs. Cyclic tensile loading, which promotes the maturation of tendon constructs in a previous work, exhibits a material-dependent effect in this study. Fibrin constructs show trending reductions in mechanical, biological, and structural properties, whereas collagen constructs only show improved tenogenic expression in the presence of mechanical stimulation. These findings highlight that components of the mechanical stimulus (e.g., strain amplitude or time of initiation) need to be tailored to the material and cell type. Given the improvements in tenogenic expression, extracellular matrix organization, and material properties during static culture, in vitro findings presented here suggest that fibrin-based constructs may be a more suitable alternative to collagen-based constructs for tissue-engineered tendon/ligament repair. PMID:25266738

  15. Microfluidic production of bioactive fibrin micro-beads embedded in crosslinked collagen used as an injectable bulking agent for urinary incontinence treatment.

    Science.gov (United States)

    Vardar, E; Larsson, H M; Allazetta, S; Engelhardt, E M; Pinnagoda, K; Vythilingam, G; Hubbell, J A; Lutolf, M P; Frey, P

    2018-02-01

    Endoscopic injection of bulking agents has been widely used to treat urinary incontinence, often due to urethral sphincter complex insufficiency. The aim of the study was to develop a novel injectable bioactive collagen-fibrin bulking agent restoring long-term continence by functional muscle tissue regeneration. Fibrin micro-beads were engineered using a droplet microfluidic system. They had an average diameter of 140 μm and recombinant fibrin-binding insulin-like growth factor-1 (α 2 PI 1-8 -MMP-IGF-1) was covalently conjugated to the beads. A plasmin fibrin degradation assay showed that 72.5% of the initial amount of α 2 PI 1-8 -MMP-IGF-1 loaded into the micro-beads was retained within the fibrin micro-beads. In vitro, the growth factor modified fibrin micro-beads enhanced cell attachment and the migration of human urinary tract smooth muscle cells, however, no change of the cellular metabolic activity was seen. These bioactive micro-beads were mixed with genipin-crosslinked homogenized collagen, acting as a carrier. The collagen concentration, the degree of crosslinking, and the mechanical behavior of this bioactive collagen-fibrin injectable were comparable to reference samples. This novel injectable showed no burst release of the growth factor, had a positive effect on cell behavior and may therefore induce smooth muscle regeneration in vivo, necessary for the functional treatment of stress and other urinary incontinences. Urinary incontinence is involuntary urine leakage, resulting from a deficient function of the sphincter muscle complex. Yet there is no functional cure for this devastating condition using current treatment options. Applied physical and surgical therapies have limited success. In this study, a novel bioactive injectable bulking agent, triggering new muscle regeneration at the injection site, has been evaluated. This injectable consists of cross-linked collagen and fibrin micro-beads, functionalized with bound insulin-like growth factor

  16. Role of Nuclear Matrix in Estrogen Regulated Gene Expression in Human Breast Cancer Cells

    Science.gov (United States)

    1998-08-01

    reticular pattern evenly distributed throughout the nucleus, excluding the nucleolus (Figure 4A). This is not so for T47D cells where a composite pattern...acetylation is required to maintain the unfolded nucleosome structure associated with transcribing DNA. Journal of Biological Chemistry 273:14516...nuclear matrix include ER, YY1, AML-1, Spl, Oct1, mutant p53, and Rb [25,28,31,34-40]. Appendix A, part 4 reviews alterations in nuclear matrix composition

  17. Fibrin Sealants in Dura Sealing: A Systematic Literature Review.

    Directory of Open Access Journals (Sweden)

    Felice Esposito

    Full Text Available Fibrin sealants are widely used in neurosurgery to seal the suture line, provide watertight closure, and prevent cerebrospinal fluid leaks. The aim of this systematic review is to summarize the current efficacy and safety literature of fibrin sealants in dura sealing and the prevention/treatment of cerebrospinal fluid leaks.A comprehensive electronic literature search was run in the following databases: Cochrane Database of Systematic Reviews, Cochrane Central Resister of Controlled Trials, clinicaltrials.gov, MEDLINE/PubMed, and EMBASE. Titles and abstracts of potential articles of interest were reviewed independently by 3 of the authors.A total of 1006 database records and additional records were identified. After screening for duplicates and relevance, a total of 78 articles were assessed by the investigators for eligibility. Thirty-eight were excluded and the full-text of 40 articles were included in the qualitative synthesis. Seven of these included only safety data and were included in the safety assessment. The remaining 33 articles included findings from 32 studies that enrolled a total of 2935 patients who were exposed to fibrin sealant. Among these 33 studies there were only 3 randomized controlled trials, with the remaining being prospective cohort analysis, case controlled studies, prospective or retrospective case series. One randomized controlled trial, with 89 patients exposed to fibrin sealant, found a greater rate of intraoperative watertight dura closure in the fibrin sealant group than the control group (92.1% versus 38.0%, p0.05. Other clinical trials evaluated the effect of fibrin sealant in the postoperative prevention of cerebrospinal fluid leaks. These were generally lower level evidence studies (ie, not prospective, randomized, controlled trials that were not designed or powered to demonstrate a significant advantage to fibrin sealant use. Two small case series studies evaluated the effect of fibrin sealants in

  18. Platelet rich fibrin in jaw defects

    Science.gov (United States)

    Nica, Diana; Ianes, Emilia; Pricop, Marius

    2016-03-01

    Platelet rich fibrin (PRF) is a tissue product of autologous origin abundant in growth factors, widely used in regenerative procedures. Aim of the study: Evaluation of the regenerative effect of PRF added in the bony defects (after tooth removal or after cystectomy) Material and methods: The comparative nonrandomized study included 22 patients divided into 2 groups. The first group (the test group) included 10 patients where the bony defects were treated without any harvesting material. The second group included 12 patients where the bony defects were filled with PRF. The bony defect design was not critical, with one to two walls missing. After the surgeries, a close clinically monitoring was carried out. The selected cases were investigated using both cone beam computer tomography (CBCT) and radiographic techniques after 10 weeks postoperatively. Results: Faster bone regeneration was observed in the bony defects filled with PRF comparing with the not grafted bony defects. Conclusions: PRF added in the bony defects accelerates the bone regeneration. This simplifies the surgical procedures and decreases the economic costs.

  19. Evaluation of Fibrin-Based Interpenetrating Polymer Networks as Potential Biomaterials for Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Olfat Gsib

    2017-12-01

    Full Text Available Interpenetrating polymer networks (IPNs have gained great attention for a number of biomedical applications due to their improved properties compared to individual components alone. In this study, we investigated the capacity of newly-developed naturally-derived IPNs as potential biomaterials for tissue engineering. These IPNs combine the biologic properties of a fibrous fibrin network polymerized at the nanoscale and the mechanical stability of polyethylene oxide (PEO. First, we assessed their cytotoxicity in vitro on L929 fibroblasts. We further evaluated their biocompatibility ex vivo with a chick embryo organotypic culture model. Subcutaneous implantations of the matrices were subsequently conducted on nude mice to investigate their biocompatibility in vivo. Our preliminary data highlighted that our biomaterials were non-cytotoxic (viability above 90%. The organotypic culture showed that the IPN matrices induced higher cell adhesion (across all the explanted organ tissues and migration (skin, intestine than the control groups, suggesting the advantages of using a biomimetic, yet mechanically-reinforced IPN-based matrix. We observed no major inflammatory response up to 12 weeks post implantation. All together, these data suggest that these fibrin-based IPNs are promising biomaterials for tissue engineering.

  20. Injectable alginate-O-carboxymethyl chitosan/nano fibrin composite hydrogels for adipose tissue engineering.

    Science.gov (United States)

    Jaikumar, Dhanya; Sajesh, K M; Soumya, S; Nimal, T R; Chennazhi, K P; Nair, Shantikumar V; Jayakumar, R

    2015-03-01

    Injectable, biodegradable scaffolds are required for soft tissue reconstruction owing to its minimally invasive approach. Such a scaffold can mimic the native extracellular matrix (ECM), provide uniform distribution of cells and overcome limitations like donor site morbidity, volume loss, etc. So, here we report two classes of biocompatible and biodegradable hydrogel blend systems namely, Alginate/O-carboxymethyl chitosan (O-CMC) and Alginate/poly (vinyl alcohol) (PVA) with the inclusion of fibrin nanoparticles in each. The hydrogels were prepared by ionic cross-linking method. The developed hydrogels were compared in terms of its swelling ratio, degradation profile, compressive strength and elastic moduli. From these preliminary findings, it was concluded that Alginate/O-CMC formed a better blend for tissue engineering applications. The potential of the formed hydrogel as an injectable scaffold was revealed by the survival of adipose derived stem cells (ADSCs) on the scaffold by its adhesion, proliferation and differentiation into adipocytes. Cell differentiation studies of fibrin incorporated hydrogel scaffolds showed better differentiation was confirmed by Oil Red O staining technique. These injectable gels have potential in soft tissue regeneration. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Evaluation of osteoblastic activity in extraction sockets treated with platelet-rich fibrin.

    Science.gov (United States)

    Baslarli, Ozgur; Tumer, Celal; Ugur, Omer; Vatankulu, Betul

    2015-01-01

    The aim of this study was to determine whether the use of platelet rich fibrin (PRF) improved the healing of extraction sockets. A total of 20 patients with bilateral soft tissue impacted mandibular third molars were included in this study. The left and right third molars were extracted during the same session. Subsequently, the PRF membrane was randomly administered to one of the extraction sockets, whereas the contralateral sockets were left without treatment. On postoperative 30. and 90. days, panoramic images and bone scintigrams were taken to evaluate the bone healing between PRF-treated and non-PRF-treated sockets. Also, periodontal evaluation was performed in the same control sessions. Dependent group t test for paired samples was used for statistical analysis. The average increase in technetium-99m methylene diphosphonate uptake as an indication of enhanced bone healing did not differ significantly between PRF-treated and non-PRF-treated sockets 30 and 90 days postoperatively. Radio-opacity that can show the bone healing on panoramic images were measured by Image J programme and they did not differ significantly. Also periodontal values did not differ significantly. PRF might not lead to enhanced bone healing in impacted mandibular third molar extraction sockets 30 and 90 days after surgery. It is thought that PRF has the potential characteristics of an autologous fibrin matrix and can accelerate the healing. To better understand the effects of PRF on healing, further research is warranted with larger sample sizes.

  2. Characteristics of an autologous leukocyte and platelet-rich fibrin patch intended for the treatment of recalcitrant wounds

    DEFF Research Database (Denmark)

    Lundquist, Rasmus; Holmstrøm, Kim; Clausen, Christian

    2012-01-01

    We have investigated the physical, biochemical, and cellular properties of an autologous leukocyte and platelet-rich fibrin patch. This was generated in an automated device from a sample of a patient's blood at the point of care. Using microscopy, cell counting, enzyme-linked immunosorbent assay...... of chronic wound fluid. By comparison with traditional platelet-rich plasma, differences in immune components were found. The relevance of these findings was assessed by showing a mitogenic and migratory effect on cultured human dermal fibroblasts. Further, we showed that fibrocytes, a cell type important......, antibody arrays, and cell culture assays, we show that the patch is a three-layered membrane comprising a fibrin sheet, a layer of platelets, and a layer of leukocytes. Mean recovery of platelets from the donated blood was 98% (±95%CI 0.8%). Mean levels of platelet-derived growth factor AB, human...

  3. Tensile strength of biological fibrin sealants: a comparative study.

    Science.gov (United States)

    Lacaze, Laurence; Le Dem, Nicolas; Bubenheim, Michael; Tsilividis, Basile; Mezghani, Julien; Schwartz, Lilian; Francois, Arnaud; Ertaud, Jean Yves; Bagot d'Arc, Maurice; Scotté, Michel

    2012-08-01

    Fibrin sealants are commonly used in liver surgery, although their effectiveness in routine clinical practice remains controversial. Individual sealant characteristics are based on hemostatic effects and adhesion properties that can be experimentally measured using the 'rat skin test' or the 'pig skin test'. This study used a more relevant and realistic experimental canine model to compare the differences in the adhesive properties of four fibrin sealants in hepatectomy: Tisseel/Tissucol, Tachosil, Quixil, and Beriplast. A partial hepatectomy was performed in beagle dogs under general anesthesia to obtain liver cross-sections. Fibrin sealants were allocated to dog livers using a Youden square design. The tensile strength measurement was performed using a traction system to measure the rupture stress point of a small wooden cylinder bonded to the liver cross-section. Significantly greater adhesion properties were observed with Tisseel/Tissucol compared with Quixil or Beriplast (P = 0.002 and 0.001, respectively). Similarly, Tachosil demonstrated significantly greater adhesive properties compared with Beriplast (P = 0.009) or Quixil (P = 0.014). No significant differences were observed between Tisseel/Tissucol and Tachosil or between Beriplast and Quixil. The results of this comparative study demonstrate that different fibrin sealants exhibit different adhesive properties. Tisseel/Tissucol and Tachosil provided greatest adhesion to liver cross-section in our canine model of hepatectomy. These results may enable the optimal choice of fibrin sealants for this procedure in clinical practice. Copyright © 2012 Elsevier Inc. All rights reserved.

  4. Fibrin(ogen)olytic activity of bumblebee venom serine protease

    International Nuclear Information System (INIS)

    Qiu Yuling; Choo, Young Moo; Yoon, Hyung Joo; Jia Jingming; Cui Zheng; Wang Dong; Kim, Doh Hoon; Sohn, Hung Dae; Jin, Byung Rae

    2011-01-01

    Bee venom is a rich source of pharmacologically active components; it has been used as an immunotherapy to treat bee venom hypersensitivity, and venom therapy has been applied as an alternative medicine. Here, we present evidence that the serine protease found in bumblebee venom exhibits fibrin(ogen)olytic activity. Compared to honeybee venom, bumblebee venom contains a higher content of serine protease, which is one of its major components. Venom serine proteases from bumblebees did not cross-react with antibodies against the honeybee venom serine protease. We provide functional evidence indicating that bumblebee (Bombus terrestris) venom serine protease (Bt-VSP) acts as a fibrin(ogen)olytic enzyme. Bt-VSP activates prothrombin and directly degrades fibrinogen into fibrin degradation products. However, Bt-VSP is not a plasminogen activator, and its fibrinolytic activity is less than that of plasmin. Taken together, our results define roles for Bt-VSP as a prothrombin activator, a thrombin-like protease, and a plasmin-like protease. These findings offer significant insight into the allergic reaction sequence that is initiated by bee venom serine protease and its potential usefulness as a clinical agent in the field of hemostasis and thrombosis. - Graphical abstract: Display Omitted Highlights: → Bumblebee venom serine protease (Bt-VSP) is a fibrin(ogen)olytic enzyme. → Bt-VSP activates prothrombin. → Bt-VSP directly degrades fibrinogen into fibrin degradation products. → Bt-VSP is a hemostatically active protein that is a potent clinical agent.

  5. Fibrin network pattern changes of platelet-rich fibrin in young versus old age group of individuals: A cell block cytology study

    Directory of Open Access Journals (Sweden)

    Shravanthi Raghav Yajamanya

    2016-01-01

    Full Text Available Background: To evaluate variations in fibrin network patterns of the platelet-rich fibrin (PRF in different age groups. Materials and Methods: Ninety-five patients were divided into three age groups: Group 1: (20–39 years; Group 2: (40–59 years; and Group 3: (60 years and above. PRF was prepared from blood samples of all patients and were subjected to cell block cytology method of histological analysis and slides were prepared to histologically assess the age-related changes in (i fibrin network patterns in terms of density and (ii entrapment of platelets and white blood cells (WBCs within fibrin meshwork. Results: Two types of fibrin network pattern arrangements noticed: Dense and loose types in three age groups. However, there was a noticeable decrease in the dense type of fibrin network with progressing age and increase in the loose type of fibrin arrangement. Furthermore, variation in a number of platelets and WBCs entrapped within fibrin network in relation to age was noticed. Conclusion: From the current study it can be concluded that age can be considered as one of the influencing factors on quality of PRF in terms of fibrin network patterns and hence, platelet and WBCs entrapment within these fibrin networks.

  6. Identification of quantitative trait loci for fibrin clot phenotypes: the EuroCLOT study

    DEFF Research Database (Denmark)

    Williams, Frances M K; Carter, Angela M; Kato, Bernet

    2009-01-01

    OBJECTIVE: Fibrin makes up the structural basis of an occlusive arterial thrombus, and variability in fibrin phenotype relates to cardiovascular risk. The aims of the current study from the EU consortium EuroCLOT were to (1) determine the heritability of fibrin phenotypes and (2) identify QTLs as...

  7. Peeping into human renal calcium oxalate stone matrix: characterization of novel proteins involved in the intricate mechanism of urolithiasis.

    Directory of Open Access Journals (Sweden)

    Kanu Priya Aggarwal

    Full Text Available BACKGROUND: The increasing number of patients suffering from urolithiasis represents one of the major challenges which nephrologists face worldwide today. For enhancing therapeutic outcomes of this disease, the pathogenic basis for the formation of renal stones is the need of hour. Proteins are found as major component in human renal stone matrix and are considered to have a potential role in crystal-membrane interaction, crystal growth and stone formation but their role in urolithiasis still remains obscure. METHODS: Proteins were isolated from the matrix of human CaOx containing kidney stones. Proteins having MW>3 kDa were subjected to anion exchange chromatography followed by molecular-sieve chromatography. The effect of these purified proteins was tested against CaOx nucleation and growth and on oxalate injured Madin-Darby Canine Kidney (MDCK renal epithelial cells for their activity. Proteins were identified by Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF MS followed by database search with MASCOT server. In silico molecular interaction studies with CaOx crystals were also investigated. RESULTS: Five proteins were identified from the matrix of calcium oxalate kidney stones by MALDI-TOF MS followed by database search with MASCOT server with the competence to control the stone formation process. Out of which two proteins were promoters, two were inhibitors and one protein had a dual activity of both inhibition and promotion towards CaOx nucleation and growth. Further molecular modelling calculations revealed the mode of interaction of these proteins with CaOx at the molecular level. CONCLUSIONS: We identified and characterized Ethanolamine-phosphate cytidylyltransferase, Ras GTPase-activating-like protein, UDP-glucose:glycoprotein glucosyltransferase 2, RIMS-binding protein 3A, Macrophage-capping protein as novel proteins from the matrix of human calcium oxalate stone which play a critical role in kidney stone

  8. Human macrophages primed with angiogenic factors show dynamic plasticity, irrespective of extracellular matrix components

    NARCIS (Netherlands)

    Ploeger, Diana T. A.; van Putten, Sander M.; Koerts, Jasper A.; van Luyn, Marja J. A.; Harmsen, Martin C.

    Macrophages are important in inflammation as well as in tissue repair processes. They can be activated by various stimuli and classified into two major groups: M1 (classically activated) or M2 (alternatively activated). Inflammation, angiogenesis and matrix remodeling play a major role in tissue

  9. Characterisation of cell cycle arrest and terminal differentiation in a maximally proliferative human epithelial tissue: Lessons from the human hair follicle matrix.

    Science.gov (United States)

    Purba, Talveen S; Brunken, Lars; Peake, Michael; Shahmalak, Asim; Chaves, Asuncion; Poblet, Enrique; Ceballos, Laura; Gandarillas, Alberto; Paus, Ralf

    2017-09-01

    Human hair follicle (HF) growth and hair shaft formation require terminal differentiation-associated cell cycle arrest of highly proliferative matrix keratinocytes. However, the regulation of this complex event remains unknown. CIP/KIP family member proteins (p21 CIP1 , p27 KIP1 and p57 KIP2 ) regulate cell cycle progression/arrest, endoreplication, differentiation and apoptosis. Since they have not yet been adequately characterized in the human HF, we asked whether and where CIP/KIP proteins localise in the human hair matrix and pre-cortex in relation to cell cycle activity and HF-specific epithelial cell differentiation that is marked by keratin 85 (K85) protein expression. K85 expression coincided with loss or reduction in cell cycle activity markers, including in situ DNA synthesis (EdU incorporation), Ki-67, phospho-histone H3 and cyclins A and B1, affirming a post-mitotic state of pre-cortical HF keratinocytes. Expression of CIP/KIP proteins was found abundantly within the proliferative hair matrix, concomitant with a role in cell cycle checkpoint control. p21 CIP1 , p27 KIP1 and cyclin E persisted within post-mitotic keratinocytes of the pre-cortex, whereas p57 KIP2 protein decreased but became nuclear. These data imply a supportive role for CIP/KIP proteins in maintaining proliferative arrest, differentiation and anti-apoptotic pathways, promoting continuous hair bulb growth and hair shaft formation in anagen VI. Moreover, post-mitotic hair matrix regions contained cells with enlarged nuclei, and DNA in situ hybridisation showed cells that were >2N in the pre-cortex. This suggests that CIP/KIP proteins might counterbalance cyclin E to control further rounds of DNA replication in a cell population that has a propensity to become tetraploid. These data shed new light on the in situ-biography of human hair matrix keratinocytes on their path of active cell cycling, arrest and terminal differentiation, and showcase the human HF as an excellent, clinically

  10. The effects of collagen-rich extracellular matrix on the intracellular delivery of glycol chitosan nanoparticles in human lung fibroblasts.

    Science.gov (United States)

    Yhee, Ji Young; Yoon, Hong Yeol; Kim, Hyunjoon; Jeon, Sangmin; Hergert, Polla; Im, Jintaek; Panyam, Jayanth; Kim, Kwangmeyung; Nho, Richard Seonghun

    2017-01-01

    Recent progress in nanomedicine has shown a strong possibility of targeted therapy for obstinate chronic lung diseases including idiopathic pulmonary fibrosis (IPF). IPF is a fatal lung disease characterized by persistent fibrotic fibroblasts in response to type I collagen-rich extracellular matrix. As a pathological microenvironment is important in understanding the biological behavior of nanoparticles, in vitro cellular uptake of glycol chitosan nanoparticles (CNPs) in human lung fibroblasts was comparatively studied in the presence or absence of type I collagen matrix. Primary human lung fibroblasts from non-IPF and IPF patients (n=6/group) showed significantly increased cellular uptake of CNPs (>33.6-78.1 times) when they were cultured on collagen matrix. To elucidate the underlying mechanism of enhanced cellular delivery of CNPs in lung fibroblasts on collagen, cells were pretreated with chlorpromazine, genistein, and amiloride to inhibit clathrin-mediated endocytosis, caveolae-mediated endocytosis, and macropinocytosis, respectively. Amiloride pretreatment remarkably reduced the cellular uptake of CNPs, suggesting that lung fibroblasts mainly utilize the macropinocytosis-dependent mechanism when interacted with collagen. In addition, the internalization of CNPs was predominantly suppressed by a phosphoinositide 3-kinase (PI3K) inhibitor in IPF fibroblasts, indicating that enhanced PI3K activity associated with late-stage macropinocytosis can be particularly important for the enhanced cellular delivery of CNPs in IPF fibroblasts. Our study strongly supports the concept that a pathological microenvironment which surrounds lung fibroblasts has a significant impact on the intracellular delivery of nanoparticles. Based on the property of enhanced intracellular delivery of CNPs when fibroblasts are made to interact with a collagen-rich matrix, we suggest that CNPs may have great potential as a drug-carrier system for targeting fibrotic lung fibroblasts.

  11. Fibrinous pericarditis secondary to bacterial infection in a cat.

    Science.gov (United States)

    Tagawa, Michihito; Kurashima, Chihiro; Shimbo, Genya; Omura, Hiroshi; Koyama, Kenji; Horiuchi, Noriyuki; Kobayashi, Yoshiyasu; Kawamoto, Keiko; Miyahara, Kazuro

    2017-06-10

    A three-year-old spayed domestic short-haired cat presented for evaluation of weight loss, cardiomegaly and pleural effusion. Echocardiographic examination demonstrated a thickened pericardium with mild pericardial effusion and a large volume of pleural effusion characterized by exudate. Although the cat was treated with antibiotics, the clinical symptoms did not improve. The cat developed dyspnea and died on day 7. Necropsy revealed a large amount of modified transudates ascites, pleural effusion and markedly dilated pericardium. Histopathological examination revealed severe exudation of fibrin and granulation tissue in a thick layer of the epicardium. The cat was diagnosed with fibrinous pericarditis secondary to bacterial infection.

  12. Hyaluronan and hyaluronectin in the extracellular matrix of human brain tumour stroma.

    Science.gov (United States)

    Delpech, B; Maingonnat, C; Girard, N; Chauzy, C; Maunoury, R; Olivier, A; Tayot, J; Creissard, P

    1993-01-01

    Hyaluronan (HA) and the hyaluronan-binding glycoprotein hyaluronectin (HN) were measured in 23 gliomas and 8 meningiomas and their location was revisited in 35 tumours. A clear-cut difference was found in the HN/HA ratio values of glioblastomas (below 0.5) and that of astrocytomas (above 0.5 P edification of the extracellular matrix. In meningiomas only the stroma would be responsible for HA and HN production.

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

    Science.gov (United States)

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

    1983-01-01

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

  14. Repeated short climatic change affects the epidermal differentiation program and leads to matrix remodeling in a human organotypic skin model.

    Science.gov (United States)

    Boutrand, Laetitia-Barbollat; Thépot, Amélie; Muther, Charlotte; Boher, Aurélie; Robic, Julie; Guéré, Christelle; Vié, Katell; Damour, Odile; Lamartine, Jérôme

    2017-01-01

    Human skin is subject to frequent changes in ambient temperature and humidity and needs to cope with these environmental modifications. To decipher the molecular response of human skin to repeated climatic change, a versatile model of skin equivalent subject to "hot-wet" (40°C, 80% relative humidity [RH]) or "cold-dry" (10°C, 40% RH) climatic stress repeated daily was used. To obtain an exhaustive view of the molecular mechanisms elicited by climatic change, large-scale gene expression DNA microarray analysis was performed and modulated function was determined by bioinformatic annotation. This analysis revealed several functions, including epidermal differentiation and extracellular matrix, impacted by repeated variations in climatic conditions. Some of these molecular changes were confirmed by histological examination and protein expression. Both treatments (hot-wet and cold-dry) reduced the expression of genes encoding collagens, laminin, and proteoglycans, suggesting a profound remodeling of the extracellular matrix. Strong induction of the entire family of late cornified envelope genes after cold-dry exposure, confirmed at protein level, was also observed. These changes correlated with an increase in epidermal differentiation markers such as corneodesmosin and a thickening of the stratum corneum, indicating possible implementation of defense mechanisms against dehydration. This study for the first time reveals the complex pattern of molecular response allowing adaption of human skin to repeated change in its climatic environment.

  15. Broccoli and watercress suppress matrix metalloproteinase-9 activity and invasiveness of human MDA-MB-231 breast cancer cells

    International Nuclear Information System (INIS)

    Rose, Peter; Huang, Qing; Ong, Choon Nam; Whiteman, Matt

    2005-01-01

    A high dietary intake of cruciferous vegetables has been associated with a reduction in numerous human pathologies particularly cancer. In the current study, we examined the inhibitory effects of broccoli (Brassica oleracea var. italica) and watercress (Rorripa nasturtium aquaticum) extracts on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced cancer cell invasion and matrix metalloproteinase-9 activity using human MDA-MB-231 breast cancer cells. Aberrant overexpression of matrix metalloproteinases, including metalloproteinase-9, is associated with increased invasive potential in cancer cell lines. Our results demonstrate that extracts of broccoli and Rorripa suppressed TPA-induced MMP-9 activity and invasiveness in a concentration dependant manner as determined by zymographic analysis. Furthermore, fractionation of individual extracts followed by liquid chromatography mass spectroscopy analysis (LC-MS) revealed that the inhibitory effects of each vegetable were associated with the presence of 4-methysulfinylbutyl (sulforaphane) and 7-methylsulphinylheptyl isothiocyanates. Taken together, our data indicate that isothiocyanates derived form broccoli and Rorripa inhibit metalloproteinase 9 activities and also suppress the invasive potential of human MDA-MB-231 breast cancer cells in vitro. The inhibitory effects observed in the current study may contribute to the suppression of carcinogenesis by diets high in cruciferous vegetables

  16. Extracellular Matrix-Mediated Maturation of Human Pluripotent Stem Cell-Derived Cardiac Monolayer Structure and Electrophysiological Function.

    Science.gov (United States)

    Herron, Todd J; Rocha, Andre Monteiro Da; Campbell, Katherine F; Ponce-Balbuena, Daniela; Willis, B Cicero; Guerrero-Serna, Guadalupe; Liu, Qinghua; Klos, Matt; Musa, Hassan; Zarzoso, Manuel; Bizy, Alexandra; Furness, Jamie; Anumonwo, Justus; Mironov, Sergey; Jalife, José

    2016-04-01

    Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) monolayers generated to date display an immature embryonic-like functional and structural phenotype that limits their utility for research and cardiac regeneration. In particular, the electrophysiological function of hPSC-CM monolayers and bioengineered constructs used to date are characterized by slow electric impulse propagation velocity and immature action potential profiles. Here, we have identified an optimal extracellular matrix for significant electrophysiological and structural maturation of hPSC-CM monolayers. hPSC-CM plated in the optimal extracellular matrix combination have impulse propagation velocities ≈2× faster than previously reported (43.6±7.0 cm/s; n=9) and have mature cardiomyocyte action potential profiles, including hyperpolarized diastolic potential and rapid action potential upstroke velocity (146.5±17.7 V/s; n=5 monolayers). In addition, the optimal extracellular matrix promoted hypertrophic growth of cardiomyocytes and the expression of key mature sarcolemmal (SCN5A, Kir2.1, and connexin43) and myofilament markers (cardiac troponin I). The maturation process reported here relies on activation of integrin signaling pathways: neutralization of β1 integrin receptors via blocking antibodies and pharmacological blockade of focal adhesion kinase activation prevented structural maturation. Maturation of human stem cell-derived cardiomyocyte monolayers is achieved in a 1-week period by plating cardiomyocytes on PDMS (polydimethylsiloxane) coverslips rather than on conventional 2-dimensional cell culture formats, such as glass coverslips or plastic dishes. Activation of integrin signaling and focal adhesion kinase is essential for significant maturation of human cardiac monolayers. © 2016 American Heart Association, Inc.

  17. Platelet-rich fibrin prepared from stored whole-blood samples.

    Science.gov (United States)

    Isobe, Kazushige; Suzuki, Masashi; Watanabe, Taisuke; Kitamura, Yutaka; Suzuki, Taiji; Kawabata, Hideo; Nakamura, Masayuki; Okudera, Toshimitsu; Okudera, Hajime; Uematsu, Kohya; Nakata, Koh; Tanaka, Takaaki; Kawase, Tomoyuki

    2017-12-01

    In regenerative therapy, self-clotted platelet concentrates, such as platelet-rich fibrin (PRF), are generally prepared on-site and are immediately used for treatment. If blood samples or prepared clots can be preserved for several days, their clinical applicability will expand. Here, we prepared PRF from stored whole-blood samples and examined their characteristics. Blood samples were collected from non-smoking, healthy male donors (aged 27-67 years, N = 6), and PRF clots were prepared immediately or after storage for 1-2 days. Fibrin fiber was examined by scanning electron microscopy. Bioactivity was evaluated by means of a bioassay system involving human periosteal cells, whereas PDGF-BB concentrations were determined by an enzyme-linked immunosorbent assay. Addition of optimal amounts of a 10% CaCl 2 solution restored the coagulative ability of whole-blood samples that contained an anticoagulant (acid citrate dextrose) and were stored for up to 2 days at ambient temperature. In PRF clots prepared from the stored whole-blood samples, the thickness and cross-links of fibrin fibers were almost identical to those of freshly prepared PRF clots. PDGF-BB concentrations in the PRF extract were significantly lower in stored whole-blood samples than in fresh samples; however, both extracts had similar stimulatory effects on periosteal-cell proliferation. Quality of PRF clots prepared from stored whole-blood samples is not reduced significantly and can be ensured for use in regenerative therapy. Therefore, the proposed method enables a more flexible treatment schedule and choice of a more suitable platelet concentrate immediately before treatment, not after blood collection.

  18. Three-dimensional structure of the human immunodeficiency virus type 1 matrix protein.

    Science.gov (United States)

    Massiah, M A; Starich, M R; Paschall, C; Summers, M F; Christensen, A M; Sundquist, W I

    1994-11-25

    The HIV-1 matrix protein forms an icosahedral shell associated with the inner membrane of the mature virus. Genetic analyses have indicated that the protein performs important functions throughout the viral life-cycle, including anchoring the transmembrane envelope protein on the surface of the virus, assisting in viral penetration, transporting the proviral integration complex across the nuclear envelope, and localizing the assembling virion to the cell membrane. We now report the three-dimensional structure of recombinant HIV-1 matrix protein, determined at high resolution by nuclear magnetic resonance (NMR) methods. The HIV-1 matrix protein is the first retroviral matrix protein to be characterized structurally and only the fourth HIV-1 protein of known structure. NMR signal assignments required recently developed triple-resonance (1H, 13C, 15N) NMR methodologies because signals for 91% of 132 assigned H alpha protons and 74% of the 129 assignable backbone amide protons resonate within chemical shift ranges of 0.8 p.p.m. and 1 p.p.m., respectively. A total of 636 nuclear Overhauser effect-derived distance restraints were employed for distance geometry-based structure calculations, affording an average of 13.0 NMR-derived distance restraints per residue for the experimentally constrained amino acids. An ensemble of 25 refined distance geometry structures with penalties (sum of the squares of the distance violations) of 0.32 A2 or less and individual distance violations under 0.06 A was generated; best-fit superposition of ordered backbone heavy atoms relative to mean atom positions afforded root-mean-square deviations of 0.50 (+/- 0.08) A. The folded HIV-1 matrix protein structure is composed of five alpha-helices, a short 3(10) helical stretch, and a three-strand mixed beta-sheet. Helices I to III and the 3(10) helix pack about a central helix (IV) to form a compact globular domain that is capped by the beta-sheet. The C-terminal helix (helix V) projects away

  19. Human lung fibroblast-derived matrix facilitates vascular morphogenesis in 3D environment and enhances skin wound healing.

    Science.gov (United States)

    Du, Ping; Suhaeri, Muhammad; Ha, Sang Su; Oh, Seung Ja; Kim, Sang-Heon; Park, Kwideok

    2017-05-01

    Extracellular matrix (ECM) is crucial to many aspects of vascular morphogenesis and maintenance of vasculature function. Currently the recapitulation of angiogenic ECM microenvironment is still challenging, due mainly to its diverse components and complex organization. Here we investigate the angiogenic potential of human lung fibroblast-derived matrix (hFDM) in creating a three-dimensional (3D) vascular construct. hFDM was obtained via decellularization of in vitro cultured human lung fibroblasts and analyzed via immunofluorescence staining and ELISA, which detect multiple ECM macromolecules and angiogenic growth factors (GFs). Human umbilical vein endothelial cells (HUVECs) morphology was more elongated and better proliferative on hFDM than on gelatin-coated substrate. To prepare 3D construct, hFDM is collected, quantitatively analyzed, and incorporated in collagen hydrogel (Col) with HUVECs. Capillary-like structure (CLS) formation at 7day was significantly better with the groups containing higher doses of hFDM compared to the Col group (control). Moreover, the group (Col/hFDM/GFs) with both hFDM and angiogenic GFs (VEGF, bFGF, SDF-1) showed the synergistic activity on CLS formation and found much larger capillary lumen diameters with time. Further analysis of hFDM via angiogenesis antibody array kit reveals abundant biochemical cues, such as angiogenesis-related cytokines, GFs, and proteolytic enzymes. Significantly up-regulated expression of VE-cadherin and ECM-specific integrin subunits was also noticed in Col/hFDM/GFs. In addition, transplantation of Col/hFMD/GFs with HUVECs in skin wound model presents more effective re-epithelialization, many regenerated hair follicles, better transplanted cells viability, and advanced neovascularization. We believe that current system is a very promising platform for 3D vasculature construction in vitro and for cell delivery toward therapeutic applications in vivo. Functional 3D vasculature construction in vitro is still

  20. Two-way regulation between cells and aligned collagen fibrils: local 3D matrix formation and accelerated neural differentiation of human decidua parietalis placental stem cells.

    Science.gov (United States)

    Li, Wen; Zhu, Bofan; Strakova, Zuzana; Wang, Rong

    2014-08-08

    It has been well established that an aligned matrix provides structural and signaling cues to guide cell polarization and cell fate decision. However, the modulation role of cells in matrix remodeling and the feedforward effect on stem cell differentiation have not been studied extensively. In this study, we report on the concerted changes of human decidua parietalis placental stem cells (hdpPSCs) and the highly ordered collagen fibril matrix in response to cell-matrix interaction. With high-resolution imaging, we found the hdpPSCs interacted with the matrix by deforming the cell shape, harvesting the nearby collagen fibrils, and reorganizing the fibrils around the cell body to transform a 2D matrix to a localized 3D matrix. Such a unique 3D matrix prompted high expression of β-1 integrin around the cell body that mediates and facilitates the stem cell differentiation toward neural cells. The study offers insights into the coordinated, dynamic changes at the cell-matrix interface and elucidates cell modulation of its matrix to establish structural and biochemical cues for effective cell growth and differentiation. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. The mechanical properties of human adipose tissues and their relationships to the structure and composition of the extracellular matrix.

    Science.gov (United States)

    Alkhouli, Nadia; Mansfield, Jessica; Green, Ellen; Bell, James; Knight, Beatrice; Liversedge, Neil; Tham, Ji Chung; Welbourn, Richard; Shore, Angela C; Kos, Katarina; Winlove, C Peter

    2013-12-01

    Adipose tissue (AT) expansion in obesity is characterized by cellular growth and continuous extracellular matrix (ECM) remodeling with increased fibrillar collagen deposition. It is hypothesized that the matrix can inhibit cellular expansion and lipid storage. Therefore, it is important to fully characterize the ECM's biomechanical properties and its interactions with cells. In this study, we characterize and compare the mechanical properties of human subcutaneous and omental tissues, which have different physiological functions. AT was obtained from 44 subjects undergoing surgery. Force/extension and stress/relaxation data were obtained. The effects of osmotic challenge were measured to investigate the cellular contribution to tissue mechanics. Tissue structure and its response to tensile strain were determined using nonlinear microscopy. AT showed nonlinear stress/strain characteristics of up to a 30% strain. Comparing paired subcutaneous and omental samples (n = 19), the moduli were lower in subcutaneous: initial 1.6 ± 0.8 (means ± SD) and 2.9 ± 1.5 kPa (P = 0.001), final 11.7 ± 6.4 and 32 ± 15.6 kPa (P matrix fibers. These results suggest that subcutaneous AT has greater capacity for expansion and recovery from mechanical deformation than omental AT.

  2. Messenger RNA for membrane-type 2 matrix metalloproteinase, MT2-MMP, is expressed in human placenta of first trimester.

    Science.gov (United States)

    Bjørn, S F; Hastrup, N; Larsen, J F; Lund, L R; Pyke, C

    2000-01-01

    An intimately regulated cell surface activation of matrix metalloproteinases (MMPs) is believed to be of critical importance for the control of trophoblast invasion. A histological investigation of the expression and localization of three different MMPs, the membrane-type matrix metalloproteinases 1 and 2 (MT1-MMP, MT2-MMP) and matrix metalloproteinase 2 (MMP-2/gelatinase A) was performed by in situ hybridization on consecutive sections from human placentae of first trimester pregnancies. Cytokeratin immunostaining identified trophoblast cells. Both normal and tubal implantation sites were studied. We observed a high degree of coexpression of MT2-MMP, MT1-MMP and MMP-2 mRNAs in single extravillous cytotrophoblasts that had invaded the endometrium and tubal wall. Furthermore, mRNAs for all three genes were also seen in cytotrophoblasts of cell islands. In contrast to this coexpression pattern, MT2-MMP expression was absent from cell columns and decidual cells, in which signals for MT1-MMP and MMP-2 mRNAs were seen. The present data on the cellular expression of MT2-MMP mRNA in placenta extend our knowledge of the proteolytic events that take place during early pregnancy. The data suggest that MT2-MMP, capable of activating MMP-2 in vitro, is involved in the invasion of extravillous cytotrophoblast, possibly related to the physiological activation of MMP-2. Copyright 2000 Harcourt Publishers Ltd.

  3. TCDD induces dermal accumulation of keratinocyte-derived matrix metalloproteinase-10 in an organotypic model of human skin

    Energy Technology Data Exchange (ETDEWEB)

    De Abrew, K. Nadira [Molecular and Environmental Toxicology Center, University of Wisconsin—Madison, Madison, WI 53706 (United States); Thomas-Virnig, Christina L.; Rasmussen, Cathy A. [Department of Pathology, University of Wisconsin—Madison, Madison, WI 53706 (United States); Bolterstein, Elyse A. [Molecular and Environmental Toxicology Center, University of Wisconsin—Madison, Madison, WI 53706 (United States); Schlosser, Sandy J. [Department of Pathology, University of Wisconsin—Madison, Madison, WI 53706 (United States); Allen-Hoffmann, B. Lynn, E-mail: blallenh@wisc.edu [Molecular and Environmental Toxicology Center, University of Wisconsin—Madison, Madison, WI 53706 (United States); Department of Pathology, University of Wisconsin—Madison, Madison, WI 53706 (United States)

    2014-05-01

    The epidermis of skin is the first line of defense against the environment. A three dimensional model of human skin was used to investigate tissue-specific phenotypes induced by the environmental contaminant, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Continuous treatment of organotypic cultures of human keratinocytes with TCDD resulted in intracellular spaces between keratinocytes of the basal and immediately suprabasal layers as well as thinning of the basement membrane, in addition to the previously reported hyperkeratinization. These tissue remodeling events were preceded temporally by changes in expression of the extracellular matrix degrading enzyme, matrix metalloproteinase-10 (MMP-10). In organotypic cultures MMP-10 mRNA and protein were highly induced following TCDD treatment. Q-PCR and immunoblot results from TCDD-treated monolayer cultures, as well as indirect immunofluorescence and immunoblot analysis of TCDD-treated organotypic cultures, showed that MMP-10 was specifically contributed by the epidermal keratinocytes but not the dermal fibroblasts. Keratinocyte-derived MMP-10 protein accumulated over time in the dermal compartment of organotypic cultures. TCDD-induced epidermal phenotypes in organotypic cultures were attenuated by the keratinocyte-specific expression of tissue inhibitor of metalloproteinase-1, a known inhibitor of MMP-10. These studies suggest that MMP-10 and possibly other MMP-10-activated MMPs are responsible for the phenotypes exhibited in the basement membrane, the basal keratinocyte layer, and the cornified layer of TCDD-treated organotypic cultures. Our studies reveal a novel mechanism by which the epithelial–stromal microenvironment is altered in a tissue-specific manner thereby inducing structural and functional pathology in the interfollicular epidermis of human skin. - Highlights: • TCDD causes hyperkeratosis and basement membrane changes in a model of human skin. • TCDD induces MMP-10 expression in organotypic cultures

  4. From single cells to tissues: interactions between the matrix and human breast cells in real time.

    Directory of Open Access Journals (Sweden)

    Clifford Barnes

    Full Text Available Mammary gland morphogenesis involves ductal elongation, branching, and budding. All of these processes are mediated by stroma--epithelium interactions. Biomechanical factors, such as matrix stiffness, have been established as important factors in these interactions. For example, epithelial cells fail to form normal acinar structures in vitro in 3D gels that exceed the stiffness of a normal mammary gland. Additionally, heterogeneity in the spatial distribution of acini and ducts within individual collagen gels suggests that local organization of the matrix may guide morphogenesis. Here, we quantified the effects of both bulk material stiffness and local collagen fiber arrangement on epithelial morphogenesis.The formation of ducts and acini from single cells and the reorganization of the collagen fiber network were quantified using time-lapse confocal microscopy. MCF10A cells organized the surrounding collagen fibers during the first twelve hours after seeding. Collagen fiber density and alignment relative to the epithelial surface significantly increased within the first twelve hours and were a major influence in the shaping of the mammary epithelium. The addition of Matrigel to the collagen fiber network impaired cell-mediated reorganization of the matrix and increased the probability of spheroidal acini rather than branching ducts. The mechanical anisotropy created by regions of highly aligned collagen fibers facilitated elongation and branching, which was significantly correlated with fiber organization. In contrast, changes in bulk stiffness were not a strong predictor of this epithelial morphology.Localized regions of collagen fiber alignment are required for ductal elongation and branching suggesting the importance of local mechanical anisotropy in mammary epithelial morphogenesis. Similar principles may govern the morphology of branching and budding in other tissues and organs.

  5. Comparison of two fibrin glues in anastomoses and skin closure.

    Science.gov (United States)

    Park, W; Kim, W H; Lee, C H; Kim, D Y; Choi, J H; Huh, J W; Sung, H M; Kim, I S; Kweon, O K

    2002-09-01

    To control intra-operative haemorrhage, fibrin glues are preferred by many surgeons because of their biological advantages and convenience of application. Manufacturers have developed a few kinds of fibrin glues with a little difference in their composition. This study was to compare the effectiveness of two commercially available fibrin glues; Greenplast (Green Cross P. D. Company, Yongin, Korea) and Tisseel (Baxter-Immuno AG, Vienna, Austria). They were applied experimentally to several kinds of surgery in dogs - renal vessel anastomosis, partial splenectomy, intestinal anastomosis and incision skin wound - and evaluated for their haemostatic and adhesive effects. When the two glues were applied in renal vessel anastomosis, the amount of haemorrhage in artery and vein decreased significantly. They also decreased the haemorrhage in partial splenectomy. At 10 min after application of the glues to an incision skin wound, the tensile strengths developed were significantly higher than that of control. The present study indicates that two-component fibrin glues have a haemostatic effect as a mechanical barrier in renal vessel anastomosis and an adhesive effect in the early stage of incision skin wound closure, and the two glues have similar effects with no complications.

  6. Gallium67 scintigraphy in fibrinous pericarditis associated with bacterial endocarditis

    International Nuclear Information System (INIS)

    Martin, P.; Verhas, M.; Devriendt, J.; Goffin, Y.

    1982-01-01

    An 80-year-old man presented with pyrexia, progressive cardiac failure and inflammation. A diagnosis of pericarditisd associated with bacterial endocarditis was suggested from Gallium 67 scintigraphy and confirmed at autpsy. This case of fibrinous pericarditis without effusion could not be diagnosed by echography or routine cardiopulmonary scintigraphy. (orig.)

  7. Submillisecond Elastic Recoil Reveals Molecular Origins of Fibrin Fiber Mechanics

    Science.gov (United States)

    Hudson, Nathan E.; Ding, Feng; Bucay, Igal; O’Brien, E. Timothy; Gorkun, Oleg V.; Superfine, Richard; Lord, Susan T.; Dokholyan, Nikolay V.; Falvo, Michael R.

    2013-01-01

    Fibrin fibers form the structural scaffold of blood clots. Thus, their mechanical properties are of central importance to understanding hemostasis and thrombotic disease. Recent studies have revealed that fibrin fibers are elastomeric despite their high degree of molecular ordering. These results have inspired a variety of molecular models for fibrin’s elasticity, ranging from reversible protein unfolding to rubber-like elasticity. An important property that has not been explored is the timescale of elastic recoil, a parameter that is critical for fibrin’s mechanical function and places a temporal constraint on molecular models of fiber elasticity. Using high-frame-rate imaging and atomic force microscopy-based nanomanipulation, we measured the recoil dynamics of individual fibrin fibers and found that the recoil was orders of magnitude faster than anticipated from models involving protein refolding. We also performed steered discrete molecular-dynamics simulations to investigate the molecular origins of the observed recoil. Our results point to the unstructured αC regions of the otherwise structured fibrin molecule as being responsible for the elastic recoil of the fibers. PMID:23790375

  8. Comparison of bioengineered human bone construct from four sources of osteogenic cells.

    Science.gov (United States)

    Ng, Angela Min-Hwei; Saim, Aminuddin Bin; Tan, Kok-Keong; Tan, G H; Mokhtar, Sabarul Afian; Rose, Isa Mohamed; Othman, Fauziah; Idrus, Ruszymah Binti Haji

    2005-01-01

    Osteoprogenitor cells have been reported to be present in periosteum, cancellous and cortical bone, and bone marrow; but no attempt to identify the best cell source for bone tissue engineering has yet been reported. In this study, we aimed to investigate the growth and differentiation pattern of cells derived from these four sources in terms of cell doubling time and expression of osteoblast-specific markers in both monolayer cells and three-dimensional cell constructs in vitro. In parallel, human plasma derived-fibrin was evaluated for use as biomaterial when forming three-dimensional bone constructs. Our findings showed osteoprogenitor cells derived from periosteum to be most proliferative followed by cortical bone, cancellous bone, and then bone marrow aspirate. Bone-forming activity was observed in constructs formed with cells derived from periosteum, whereas calcium deposition was seen throughout the constructs formed with cells derived from cancellous and cortical bones. Although no mineralization activity was seen in constructs formed with osteoprogenitor cells derived from bone marrow, well-organized lacunae as would appear in the early phase of bone reconstruction were noted. Scanning electron microscopy evaluation showed cell proliferation throughout the fibrin matrix, suggesting the possible application of human fibrin as the bioengineered tissue scaffold at non-load-bearing sites.

  9. Demineralized bone matrix and human cancellous bone enhance fixation of porous-coated titanium implants in sheep

    DEFF Research Database (Denmark)

    Babiker, Hassan; Ding, Ming; Overgaard, Søren

    2016-01-01

    matrix (DBM), alone or in combination with allograft or commercially available human cancellous bone (CB), may replace allografts, as they have the capability of inducing new bone and improving implant fixation through enhancing bone ongrowth. The purpose of this study was to investigate the effect...... of DBM alone, DBM with CB, or allograft on the fixation of porous-coated titanium implants. DBM100 and CB produced from human tissue were included. Both materials are commercially available. DBM granules are placed in pure DBM and do not contain any other carrier. Titanium alloy implants, 10 mm long × 10...... mm diameter, were inserted bilaterally into the femoral condyles of eight skeletally mature sheep. Thus, four implants with a concentric gap of 2 mm were implanted in each sheep. The gap was filled with: (a) DBM; (b) DBM:CB at a ratio of 1:3; (c) DBM:allograft at a ratio of 1:3; or (d) allograft...

  10. ADAM12 in human liver cancers: TGF-beta-regulated expression in stellate cells is associated with matrix remodeling

    DEFF Research Database (Denmark)

    Le Pabic, Hélène; Bonnier, Dominique; Wewer, Ulla M

    2003-01-01

    "A disintegrin and metalloproteinases" (ADAMs) form a family of cell-surface glycoproteins with potential protease and cell-adhesion activities. We have investigated ADAM expression in human liver cancers and their regulation by several cytokines involved in liver injury. Using degenerative RT...... carcinomas (up to 3- and 6-fold, respectively) and liver metastases from colonic carcinomas (up to 40- and 60-fold, respectively). The up-regulation of both ADAM9 and ADAM12 was correlated with an increase in matrix metalloproteinase 2 expression and activity. In conclusion, in liver cancers ADAM9 and ADAM12......-PCR, cDNA encoding sequences for ADAM9 and ADAM12 were identified in human activated hepatic stellate cells (HSCs). Northern blot analyses showed that HSCs, but not hepatocytes, expressed transcripts for ADAM9 messenger RNA (mRNA) and both the long and short forms of ADAM12. This expression...

  11. Extracellular matrix components expression in human pluripotent stem cell-derived retinal organoids recapitulates retinogenesis in vivo and reveals an important role for IMPG1 and CD44 in the development of photoreceptors and interphotoreceptor matrix.

    Science.gov (United States)

    Felemban, Majed; Dorgau, Birthe; Hunt, Nicola Claire; Hallam, Dean; Zerti, Darin; Bauer, Roman; Ding, Yuchun; Collin, Joseph; Steel, David; Krasnogor, Natalio; Al-Aama, Jumana; Lindsay, Susan; Mellough, Carla; Lako, Majlinda

    2018-05-17

    The extracellular matrix (ECM) plays an important role in numerous processes including cellular proliferation, differentiation, migration, maturation, adhesion guidance and axonal growth. To date, there has been no detailed analysis of the ECM distribution during retinal ontogenesis in humans and the functional importance of many ECM components is poorly understood. In this study, the expression of key ECM components in adult mouse and monkey retina, developing and adult human retina and retinal organoids derived from human pluripotent stem cells was studied. Our data indicate that basement membrane ECMs (Fibronectin and Collagen IV) were expressed in Bruch's membrane and the inner limiting membrane of the developing human retina, whilst the hyalectins (Versican and Brevican), cluster of differentiation 44 (CD44), photoreceptor-specific ECMs Interphotoreceptor Matrix Proteoglycan 1 (IMPG1) and Interphotoreceptor Matrix Proteoglycan 2 (IMPG2) were detected in the developing interphotoreceptor matrix (IPM). The expression of IMPG1, Versican and Brevican in the developing IPM was conserved between human developing retina and human pluripotent stem cell-derived retinal organoids. Blocking the action of CD44 and IMPG1 in pluripotent stem cell derived retinal organoids affected the development of photoreceptors, their inner/outer segments and connecting cilia and disrupted IPM formation, with IMPG1 having an earlier and more significant impact. Together, our data suggest an important role for IMPG1 and CD44 in the development of photoreceptors and IPM formation during human retinogenesis. The expression and the role of many extracellular matrix (ECM) components during human retinal development is not fully understood. In this study, expression of key ECM components (Collagen IV, Fibronectin, Brevican, Versican, IMPG1 and IMPG2) was investigated during human retinal ontogenesis. Collagen IV and Fibronectin were expressed in Bruch's membrane; whereas Brevican, Versican

  12. Cultivation of corneal endothelial cells on a pericellular matrix prepared from human decidua-derived mesenchymal cells.

    Directory of Open Access Journals (Sweden)

    Ryohei Numata

    Full Text Available The barrier and pump functions of the corneal endothelium are essential for the maintenance of corneal transparency. Although corneal transplantation is the only current therapy for treating corneal endothelial dysfunction, the potential of tissue-engineering techniques to provide highly efficient and less invasive therapy in comparison to corneal transplantation has been highly anticipated. However, culturing human corneal endothelial cells (HCECs is technically difficult, and there is no established culture protocol. The aim of this study was to investigate the feasibility of using a pericellular matrix prepared from human decidua-derived mesenchymal cells (PCM-DM as an animal-free substrate for HCEC culture for future clinical applications. PCM-DM enhanced the adhesion of monkey CECs (MCECs via integrin, promoted cell proliferation, and suppressed apoptosis. The HCECs cultured on the PCM-DM showed a hexagonal morphology and a staining profile characteristic of Na⁺/K⁺-ATPase and ZO-1 at the plasma membrane in vivo, whereas the control HCECs showed a fibroblastic phenotype. The cell density of the cultured HCECs on the PCM-DM was significantly higher than that of the control cells. These results indicate that PCM-DM provides a feasible xeno-free matrix substrate and that it offers a viable in vitro expansion protocol for HCECs while maintaining cellular functions for use as a subsequent clinical intervention for tissue-engineered based therapy of corneal endothelial dysfunction.

  13. Altered expression of mitochondrial and extracellular matrix genes in the heart of human fetuses with chromosome 21 trisomy

    Directory of Open Access Journals (Sweden)

    Olla Carlo

    2007-08-01

    Full Text Available Abstract Background The Down syndrome phenotype has been attributed to overexpression of chromosome 21 (Hsa21 genes. However, the expression profile of Hsa21 genes in trisomic human subjects as well as their effects on genes located on different chromosomes are largely unknown. Using oligonucleotide microarrays we compared the gene expression profiles of hearts of human fetuses with and without Hsa21 trisomy. Results Approximately half of the 15,000 genes examined (87 of the 168 genes on Hsa21 were expressed in the heart at 18–22 weeks of gestation. Hsa21 gene expression was globally upregulated 1.5 fold in trisomic samples. However, not all genes were equally dysregulated and 25 genes were not upregulated at all. Genes located on other chromosomes were also significantly dysregulated. Functional class scoring and gene set enrichment analyses of 473 genes, differentially expressed between trisomic and non-trisomic hearts, revealed downregulation of genes encoding mitochondrial enzymes and upregulation of genes encoding extracellular matrix proteins. There were no significant differences between trisomic fetuses with and without heart defects. Conclusion We conclude that dosage-dependent upregulation of Hsa21 genes causes dysregulation of the genes responsible for mitochondrial function and for the extracellular matrix organization in the fetal heart of trisomic subjects. These alterations might be harbingers of the heart defects associated with Hsa21 trisomy, which could be based on elusive mechanisms involving genetic variability, environmental factors and/or stochastic events.

  14. Categorical dimensions of human odor descriptor space revealed by non-negative matrix factorization

    Energy Technology Data Exchange (ETDEWEB)

    Chennubhotla, Chakra [University of Pittsburgh School of Medicine, Pittsburgh PA; Castro, Jason [Bates College

    2013-01-01

    In contrast to most other sensory modalities, the basic perceptual dimensions of olfaction remain un- clear. Here, we use non-negative matrix factorization (NMF) - a dimensionality reduction technique - to uncover structure in a panel of odor profiles, with each odor defined as a point in multi-dimensional descriptor space. The properties of NMF are favorable for the analysis of such lexical and perceptual data, and lead to a high-dimensional account of odor space. We further provide evidence that odor di- mensions apply categorically. That is, odor space is not occupied homogenously, but rather in a discrete and intrinsically clustered manner. We discuss the potential implications of these results for the neural coding of odors, as well as for developing classifiers on larger datasets that may be useful for predicting perceptual qualities from chemical structures.

  15. Effects of scaffold composition and architecture on human nasal chondrocyte redifferentiation and cartilaginous matrix deposition

    NARCIS (Netherlands)

    Miot, Sylvie; Woodfield, T.B.F.; Daniels, Alma U.; Suetterlin, Rosemarie; Peterschmitt, Iman; Heberer, Michael; van Blitterswijk, Clemens; Riesle, J.U.; Martin, Ivan

    2005-01-01

    We investigated whether the post-expansion redifferentiation and cartilage tissue formation capacity of adult human nasal chondrocytes can be regulated by controlled modifications of scaffold composition and architecture. As a model system, we used poly(ethylene

  16. A flexible matrix-based human exposure assessment framework suitable for LCA and CAA

    DEFF Research Database (Denmark)

    Jolliet, Olivier; Ernstoff, Alexi; Huang, Lei

    2016-01-01

    are not applicable to all types of near-field chemical releases from consumer products, e.g. direct dermal application. A consistent near-and far-field framework is needed for life cycle assessment (LCA) and chemical alternative assessment (CAA) to inform mitigation of human exposure to harmful chemicals. To close......Humans can be exposed to chemicals via near-field exposure pathways (e.g. through consumer product use) and far-field exposure pathways (e.g. through environmental emissions along product life cycles). Pathways are often complex where chemicals can transfer directly from products to humans during...... use or exchange between near-and far-field compartments until sub -fractions reach humans via inhalation, ingestion or dermal uptake. Currently, however, multimedia exposure models mainly focus on far-field exposure pathways. Metrics and modeling approaches used in far-field, emission-based models...

  17. Fibrin-Targeted Magnetic Resonance Imaging Allows In Vivo Quantification of Thrombus Fibrin Content and Identifies Thrombi Amenable for Thrombolysis

    Science.gov (United States)

    Jenkins, Julia; Modarai, Bijan; Wiethoff, Andrea J.; Phinikaridou, Alkystis; Grover, Steven P.; Patel, Ashish S.; Schaeffter, Tobias; Smith, Alberto; Botnar, Rene M.

    2014-01-01

    Objective Deep venous thrombosis is a major health problem. Thrombolytic therapies are effective in recanalizing the veins and preventing post-thrombotic complications, but there is no consensus on selection criteria. The aim of this study was to investigate a fibrin-specific MRI contrast agent (EP-2104R) for the accurate quantification of thrombus’ fibrin content in vivo and for the identification of thrombus suitable for thrombolysis. Approach and Results Venous thrombosis was induced in the inferior vena cava of 8- to 10-week-old male BALB/C mice and MRI performed 2, 4, 7, 10, 14, and 21 days later. Eighteen mice were scanned at each time point pre and 2 hours post injection of EP-2104R (8.0 μmol/kg) with 12 mice at each time point used to correlate fibrin contrast uptake with thrombus’ histological stage and fibrin content. Six mice at each time point were immediately subjected to intravascular thrombolytic therapy (10 mg/kg of tissue-type plasminogen activator). Mice were imaged to assess response to lytic therapy 24 hours after thrombolytic treatment. Two mice at each time point were scanned post injection of 0.2 mmol/kg of Gd-DTPA (gadolinium with diethylenetriaminepentacetate, Magnevist, Schering AG, Berlin, Germany) for control purpose. Contrast uptake was correlated positively with the fibrin content of the thrombus measured by Western blotting (R2=0.889; PThrombus relaxation rate (R1) post contrast and the change in visualized thrombus size on late gadolinium enhancement inversion recovery MRI pre–EP-2104R and post–EP-2104R injection were the best predictors for successful thrombolysis (area under the curve, 0.989 [95% confidence interval, 0.97–1.00] and 0.994 [95% confidence interval, 0.98–1.00] respectively). Conclusions MRI with a fibrin-specific contrast agent accurately estimates thrombus fibrin content in vivo and identifies thrombi that are amenable for thrombolysis. PMID:24723557

  18. Synthesis of a cyclic fibrin-like peptide and its analysis by fast atom bombardment mass spectrometry

    International Nuclear Information System (INIS)

    Young, J.D.; Costello, C.E.; Langenhove, A. van; Haber, E.; Matsueda, G.R.

    1983-01-01

    For immunochemical purposes, a cyclic 12 peptide was synthesized to model the γ-γ-chain cross-link site in human fibrin. The model was based upon the structure proposed by Chen and Doolittle which is characterized by two reciprocating epsilon-(γ-Glu)Lys bonds between adjacent fibrin γ-chains oriented in an antiparallel manner. To achieve the antiparallel orientation of the peptide backbone, Pro and Gly were inserted at positions 6 and 7 of the linear 12-peptide: acetyl-Gly-Glu-Gln-His-His-Pro-Gly-Gly-Gly-Ala-Lys-Gly-amide. The insertions were made to facilitate a reverse turn of the peptide during the last synthetic step, which was formation of the epsilon-(γ-Glu)Lys bond between Glu at position 2 and Lys at position 11 with diphenylphosphorylazide. The resulting cyclic peptide represented half of the symmetrical cross-linked region in clotted fibrin. Following purification by HPLC, both linear and cyclic 12-peptides were analyzed by fast atom bombardment mass spectrometry. Abundant molecular protonated ions were observed for both peptides. In addition, the amino acid sequence of the linear peptide and the location of the epsilon-(γ-Glu)Lys bond in the cyclized peptide could be verified. (author)

  19. Activity and expression of urokinase-type plasminogen activator and matrix metalloproteinases in human colorectal cancer

    International Nuclear Information System (INIS)

    Kim, Tae-Dong; Song, Kyoung-Sub; Li, Ge; Choi, Hoon; Park, Hae-Duck; Lim, Kyu; Hwang, Byung-Doo; Yoon, Wan-Hee

    2006-01-01

    Matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), and urokinase-type plasminogen activator (uPA) are involved in colorectal cancer invasion and metastasis. There is still debate whether the activity of MMP-2 and MMP-9 differs between tumors located in the colon and rectum. We designed this study to determine any differences in the expression of MMP-2, MMP-9 and uPA system between colon and rectal cancer tissues. Cancer tissue samples were obtained from colon carcinoma (n = 12) and rectal carcinomas (n = 10). MMP-2 and MMP-9 levels were examined using gelatin zymography and Western blotting; their endogenous inhibitors, tissue inhibitor of metalloproteinase-2 (TIMP-2) and tissue inhibitor of metalloproteinase-1 (TIMP-1), were assessed by Western blotting. uPA, uPAR and PAI-1 were examined using enzyme-linked immunosorbent assay (ELISA). The activity of uPA was assessed by casein-plasminogen zymography. In both colon and rectal tumors, MMP-2, MMP-9 and TIMP-1 protein levels were higher than in corresponding paired normal mucosa, while TIMP-2 level in tumors was significantly lower than in normal mucosa. The enzyme activities or protein levels of MMP-2, MMP-9 and their endogenous inhibitors did not reach a statistically significant difference between colon and rectal cancer compared with their normal mucosa. In rectal tumors, there was an increased activity of uPA compared with the activity in colon tumors (P = 0.0266), however urokinase-type plasminogen activator receptor (uPAR) and plasminogen activator inhibitor-1 (PAI-1) showed no significant difference between colon and rectal cancer tissues. These findings suggest that uPA may be expressed differentially in colon and rectal cancers, however, the activities or protein levels of MMP-2, MMP-9, TIMP-1, TIMP-2, PAI-1 and uPAR are not affected by tumor location in the colon or the rectum

  20. Construction of tissue-engineered small-diameter vascular grafts in fibrin scaffolds in 30 days.

    Science.gov (United States)

    Gui, Liqiong; Boyle, Michael J; Kamin, Yishai M; Huang, Angela H; Starcher, Barry C; Miller, Cheryl A; Vishnevetsky, Michael J; Niklason, Laura E

    2014-05-01

    Tissue-engineered small-diameter vascular grafts have been developed as a promising alternative to native veins or arteries for replacement therapy. However, there is still a crucial need to improve the current approaches to render the tissue-engineered blood vessels more favorable for clinical applications. A completely biological blood vessel (3-mm inner diameter) was constructed by culturing a 50:50 mixture of bovine smooth muscle cells (SMCs) with neonatal human dermal fibroblasts in fibrin gels. After 30 days of culture under pulsatile stretching, the engineered blood vessels demonstrated an average burst pressure of 913.3±150.1 mmHg (n=6), a suture retention (53.3±15.4 g) that is suitable for implantation, and a compliance (3.1%±2.5% per 100 mmHg) that is comparable to native vessels. These engineered grafts contained circumferentially aligned collagen fibers, microfibrils and elastic fibers, and differentiated SMCs, mimicking a native artery. These promising mechanical and biochemical properties were achieved in a very short culture time of 30 days, suggesting the potential of co-culturing SMCs with fibroblasts in fibrin gels to generate functional small-diameter vascular grafts for vascular reconstruction surgery.

  1. Correlation Between Placental Matrix Metalloproteinase 9 and Tumor Necrosis Factor-α Protein Expression Throughout Gestation in Normal Human Pregnancy.

    Science.gov (United States)

    Basu, Jayasri; Agamasu, Enyonam; Bendek, Bolek; Salafia, Carolyn M; Mishra, Aruna; Lopez, Julia Vasquez; Kroes, Jessica; Dragich, Sharon Claire; Thakur, Ashley; Mikhail, Magdy

    2018-04-01

    Matrix metalloproteinases (MMPs), specifically MMP-9 plays a role in human placentation. The enzyme confers an invasive ability to cytotrophoblasts and degrades the endometrial matrix as the cells infiltrate the decidua to keep up with placental growth. Since tumor necrosis factor-α (TNF-α) can induce the synthesis of MMP-9, we investigated the patterns of changes in and correlation between placental villous MMP-9 and TNF-α expressions throughout normal human gestation. Placentas were obtained from 179 normal pregnant women who underwent elective abortion or term delivery. Chorionic villi isolated from placental samples were grouped as first, second, and third trimester (7 0/7 -13 0/7 , 13 1/7 -23 6/7 , and 37 0/7 -42 4/7 weeks, respectively). Chorionic villous TNF-α and MMP-9 proteins were assayed using enzyme immunoassay kits. There were significant differences in MMP-9 and TNF-α protein expressions among the trimester groups ( P = .001). The MMP-9 protein increased progressively with an increase in gestational age (GA), but TNF-α peaked in the second trimester. Within each trimester group, we searched for the effects of variation of GA in days on the 2 variables. A significant positive correlation between MMP-9 and GA was noted in the first trimester ( r = 0.364, P = .005). No other comparisons were significant. When GA was controlled for, partial correlation revealed a significant positive correlation between TNF-α and MMP-9 only in the second trimester ( r = 0.300, P = .018). We hypothesize that the TNF-α peak and the positive correlation between TNF-α and MMP-9 in the second trimester of normal human gestation could contribute toward a successful pregnancy outcome.

  2. Usefulness of Cross-Linked Human Acellular Dermal Matrix as an Implant for Dorsal Augmentation in Rhinoplasty.

    Science.gov (United States)

    Yang, Chae Eun; Kim, Soo Jung; Kim, Ji Hee; Lee, Ju Hee; Roh, Tai Suk; Lee, Won Jai

    2018-02-01

    Asian noses are relatively small and flat compared to Caucasians; therefore, rhinoplasty procedures often focus on dorsal augmentation and tip projection rather than reduction in the nasal framework. Various autologous and alloplastic implant materials have been used for dorsal augmentation. Recently, human acellular dermal matrices have been introduced as an implant material for dorsal augmentation, camouflaging autologous implants without an additional donor site. Here, we introduce a cross-linked human acellular dermal matrix as an implant material in augmentation rhinoplasty and share the clinical experiences. Eighteen patients who underwent augmentation rhinoplasty using acellular dermal matrix from April 2014 to November 2015 were reviewed retrospectively. Clinical outcomes and complications were assessed at the outpatient clinic during the follow-up period ranging from 8 to 38 months. Contour changes were assessed through comparison of preoperative and postoperative photographs by two independent plastic surgeons. Patient satisfaction was assessed at the outpatient clinic by six questions regarding aesthetic and functional aspects. Postoperative photographs demonstrated the height of the nasal dorsum did not decrease over time except two patients whose ADM was grafted into a subperiosteal pocket. Others who underwent supraperiosteal implantation showed acceptable maintenance of dorsal height. No major complication was reported. Overall, patient satisfaction scored 81.02 out of 100. Cross-linked human ADM has advantages of both autogenous and alloplastic materials. The surgical results remain stable without complications. Therefore, it is a suitable alternative implant material for dorsal augmentation in rhinoplasty. This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

  3. Seeding for sirtuins: microseed matrix seeding to obtain crystals of human Sirt3 and Sirt2 suitable for soaking

    Energy Technology Data Exchange (ETDEWEB)

    Rumpf, Tobias [Albert-Ludwigs-University Freiburg, Albertstrasse 25, 79104 Freiburg, Baden-Württemberg (Germany); Gerhardt, Stefan; Einsle, Oliver, E-mail: einsle@biochemie.uni-freiburg.de [Albert-Ludwigs-University Freiburg, Albertstrasse 21, 79104 Freiburg, Baden-Württemberg (Germany); Jung, Manfred, E-mail: einsle@biochemie.uni-freiburg.de [Albert-Ludwigs-University Freiburg, Albertstrasse 25, 79104 Freiburg, Baden-Württemberg (Germany)

    2015-11-18

    In the present study, microseed matrix seeding was successfully applied to obtain a large number of crystals of the human sirtuin isotypes Sirt2 and Sirt3. These crystals appeared predictably in diverse crystallization conditions, diffracted to a higher resolution than reported in the literature and were subsequently used to study the protein–ligand interactions of two indole inhibitors. Sirtuins constitute a family of NAD{sup +}-dependent enzymes that catalyse the cleavage of various acyl groups from the ∊-amino group of lysines. They regulate a series of cellular processes and their misregulation has been implicated in various diseases, making sirtuins attractive drug targets. To date, only a few sirtuin modulators have been reported that are suitable for cellular research and their development has been hampered by a lack of structural information. In this work, microseed matrix seeding (MMS) was used to obtain crystals of human Sirt3 in its apo form and of human Sirt2 in complex with ADP ribose (ADPR). Crystal formation using MMS was predictable, less error-prone and yielded a higher number of crystals per drop than using conventional crystallization screening methods. The crystals were used to solve the crystal structures of apo Sirt3 and of Sirt2 in complex with ADPR at an improved resolution, as well as the crystal structures of Sirt2 in complex with ADPR and the indoles EX527 and CHIC35. These Sirt2–ADPR–indole complexes unexpectedly contain two indole molecules and provide novel insights into selective Sirt2 inhibition. The MMS approach for Sirt2 and Sirt3 may be used as the basis for structure-based optimization of Sirt2/3 inhibitors in the future.

  4. Human aqueous humor levels of transforming growth factor-β2: Association with matrix metalloproteinases/tissue inhibitors of matrix metalloproteinases

    OpenAIRE

    Jia, Yan; Yue, Yu; Hu, Dan-Ning; Chen, Ji-Li; Zhou, Ji-Bo

    2017-01-01

    The present study aims to investigate the association of transforming growth factor-β2 (TGF-β2) and matrix metalloproteinases (MMPs), MMP-2 and MMP-3, and tissue inhibitors of matrix metalloproteinases (TIMPs), TIMP-1, TIMP-2 and TIMP-3 in the aqueous humor of patients with high myopia or cataracts. The levels of TGF-β2 and MMPs/TIMPs were measured with the Luminex xMAP Technology using commercially available Milliplex xMAP kits. The association between TGF-β2 and MMPs/TIMPs levels was analyz...

  5. Novel aspects of the pathogenesis of aneurysms of the abdominal aorta in humans

    DEFF Research Database (Denmark)

    Michel, Jean-Baptiste; Martin-Ventura, José-Luis; Egido, Jesus

    2011-01-01

    and the adventitial response in the development of human AAA. The intraluminal thrombus exerts its pathogenic effect through platelet activation, fibrin formation, binding of plasminogen and its activators, and trapping of erythrocytes and neutrophils, leading to oxidative and proteolytic injury of the arterial wall...... arterial wall rupture. The roles of matrix metalloproteinases and plasmin generation in progression of AAA have been demonstrated both in animal models and in clinical studies. In the present review, we highlight recent studies addressing the role of the haemoglobin-rich, intraluminal thrombus...

  6. Clinicopathological correlation of keratinocyte growth factor and matrix metalloproteinase-9 expression in human gastric cancer.

    Science.gov (United States)

    Zhang, Qing; Wang, Ping; Shao, Ming; Chen, Shi-Wen; Xu, Zhi-Feng; Xu, Feng; Yang, Zhong-Yin; Liu, Bing-Ya; Gu, Qin-Long; Zhang, Wen-Jian; Li, Yong

    2015-01-01

    Keratinocyte growth factor (KGF) is reported to be implicated in the growth of some cancer cells. Matrix metalloproteinase 9 (MMP-9) is thought to enhance the tumor invasion and metastasis ability. This study was aimed at analyzing the relationship between KGF and MMP-9 expression and patients' clinicopathological characteristics to clarify the clinical significance of the expression of KGF and MMP-9 in gastric cancer. Tissue samples from 161 patients with primary gastric cancer were investigated using immunohistochemistry. The relationship between KGF and/or MMP-9 expression and clinicopathological characteristics was analyzed. KGF expression and MMP-9 expression in gastric cancer tissue were observed in 62 cases (38.5%) and 97 cases (60.2%), respectively. MMP-9 was significantly associated with depth of invasion, lymph node metastasis and TNM stage. The prognosis of MMP-9-positive patients was significantly poorer than that of MMP-9-negative patients (p = 0.009). KGF expression was positively correlated with MMP-9 expression in gastric cancer, and the prognosis of patients with both KGF- and MMP-9-positive tumors was significantly worse than that of patients with negative tumors for either factor (p = 0.045). Expression of MMP-9 was revealed to be an independent prognostic factor (p = 0.026). Coexpression of KGF and MMP-9 in gastric cancer could be a useful prognostic factor, and MMP-9 might also serve as a novel target for both prognostic prediction and therapeutics.

  7. Expression of matrix metalloproteinases 2 and 9 in human gastric cancer and superficial gastritis.

    Science.gov (United States)

    Sampieri, Clara Luz; de la Peña, Sol; Ochoa-Lara, Mariana; Zenteno-Cuevas, Roberto; León-Córdoba, Kenneth

    2010-03-28

    To assess expression of matrix metalloproteinases 2 (MMP2) and MMP9 in gastric cancer, superficial gastritis and normal mucosa, and to measure metalloproteinase activity. MMP2 and MMP9 mRNA expression was determined by quantitative real-time polymerase chain reaction. Normalization was carried out using three different factors. Proteins were analyzed by quantitative gelatin zymography (qGZ). 18S ribosomal RNA (18SRNA) was very highly expressed, while hypoxanthine ribosyltransferase-1 (HPRT-1) was moderately expressed. MMP2 was highly expressed, while MMP9 was not detected or lowly expressed in normal tissues, moderately or highly expressed in gastritis and highly expressed in cancer. Relative expression of 18SRNA and HPRT-1 showed no significant differences. Significant differences in MMP2 and MMP9 were found between cancer and normal tissue, but not between gastritis and normal tissue. Absolute quantification of MMP9 echoed this pattern, but differential expression of MMP2 proved conflictive. Analysis by qGZ indicated significant differences between cancer and normal tissue in MMP-2, total MMP-9, 250 and 110 kDa bands. MMP9 expression is enhanced in gastric cancer compared to normal mucosa; interpretation of differential expression of MMP2 is difficult to establish.

  8. Heparin modulates human intestinal smooth muscle (HISM) cell proliferation and matrix production

    International Nuclear Information System (INIS)

    Graham, M.; Perr, H.; Drucker, D.E.; Diegelmann, R.F.

    1986-01-01

    (HISM) cell proliferation and collagen production may play a role in the pathogenesis of intestinal stricture in Crohn's disease. The present studies were performed to evaluate the effects of heparin, a known modulator of vascular smooth muscle cells, on HISM cell proliferation and collagen production. Heparin (100 μg/ml) was added daily to HISM cell cultures for cell proliferation studies and for 24 hours at various time points during culture for collagen synthesis studies. Collagen synthesis was determined by the uptake of 3 H proline into collagenase-sensitive protein. Heparin completely inhibited cell proliferation for 7 days, after which cell numbers increased but at a slower rate than controls. Cells released from heparin inhibition demonstrated catch-up growth to control levels. Collagen production was significantly inhibited by 24 hours exposure to heparin but only at those times during culture when collagen synthesis was maximal (8 to 12 days). Non-collagen protein synthesis was inhibited by heparin at all time points during culture. Heparin through its modulation of HISM cells may play an important role in the control of the extracellular matrix of the intestinal wall

  9. Imaging-guided two-photon excitation-emission-matrix measurements of human skin tissues

    Science.gov (United States)

    Yu, Yingqiu; Lee, Anthony M. D.; Wang, Hequn; Tang, Shuo; Zhao, Jianhua; Lui, Harvey; Zeng, Haishan

    2012-07-01

    There are increased interests on using multiphoton imaging and spectroscopy for skin tissue characterization and diagnosis. However, most studies have been done with just a few excitation wavelengths. Our objective is to perform a systematic study of the two-photon fluorescence (TPF) properties of skin fluorophores, normal skin, and diseased skin tissues. A nonlinear excitation-emission-matrix (EEM) spectroscopy system with multiphoton imaging guidance was constructed. A tunable femtosecond laser was used to vary excitation wavelengths from 730 to 920 nm for EEM data acquisition. EEM measurements were performed on excised fresh normal skin tissues, seborrheic keratosis tissue samples, and skin fluorophores including: NADH, FAD, keratin, melanin, collagen, and elastin. We found that in the stratum corneum and upper epidermis of normal skin, the cells have large sizes and the TPF originates from keratin. In the lower epidermis, cells are smaller and TPF is dominated by NADH contributions. In the dermis, TPF is dominated by elastin components. The depth resolved EEM measurements also demonstrated that keratin structure has intruded into the middle sublayers of the epidermal part of the seborrheic keratosis lesion. These results suggest that the imaging guided TPF EEM spectroscopy provides useful information for the development of multiphoton clinical devices for skin disease diagnosis.

  10. Organic composite-mediated surface coating of human acellular bone matrix with strontium.

    Science.gov (United States)

    Huang, Yi-Zhou; Wang, Jing-Jing; Huang, Yong-Can; Wu, Cheng-Guang; Zhang, Yi; Zhang, Chao-Liang; Bai, Lin; Xie, Hui-Qi; Li, Zhao-Yang; Deng, Li

    2018-03-01

    Acellular bone matrix (ACBM) provides an osteoconductive scaffold for bone repair, but its osteoinductivity is poor. Strontium (Sr) improves the osteoinductivity of bone implants. In this study, we developed an organic composite-mediated strontium coating strategy for ACBM scaffolds by using the ion chelating ability of carboxymethyl cellulose (CMC) and the surface adhesion ability of dopamine (DOPA). The organic coating composite, termed the CMC-DOPA-Sr composite, was synthesized under a mild condition, and its chemical structure and strontium ion chelating ability were then determined. After surface decoration, the physicochemical properties of the strontium-coated ACBM (ACBM-Sr) scaffolds were characterized, and their biocompatibility and osteoinductivity were determined in vitro and in vivo. The results showed that the CMC-DOPA-Sr composite facilitated strontium coating on the surface of ACBM scaffolds. The ACBM-Sr scaffolds possessed a sustained strontium ion release profile, exhibited good cytocompatibility, and enhanced the osteogenic differentiation of mesenchymal stem cells in vitro. Furthermore, the ACBM-Sr scaffolds showed good histocompatibility after subcutaneous implantation in nude mice. Taken together, this study provided a simple and mild strategy to realize strontium coating for ACBM scaffolds, which resulted in good biocompatibility and improved osteoinductivity. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. GH/IGF-I axis and matrix adaptation of the musculotendinous tissue to exercise in humans

    DEFF Research Database (Denmark)

    Heinemeier, K M; Mackey, Abigail; Doessing, S

    2012-01-01

    cells (satellite cells), as increased satellite cell numbers are found in human muscle with increased GH/IGF-I levels, despite no change in myofibrillar protein synthesis. Although advanced age is associated with both a reduction in the GH/IGF-I axis activity, and in skeletal muscle mass (sarcopenia...

  12. Correlation of expression and activity of matrix metalloproteinase-9 and -2 in human gingival cells of periodontitis patients.

    Science.gov (United States)

    Kim, Kyung-A; Chung, Soo-Bong; Hawng, Eun-Young; Noh, Seung-Hyun; Song, Kwon-Ho; Kim, Hanna-Hyun; Kim, Cheorl-Ho; Park, Young-Guk

    2013-02-01

    Matrix metalloproteinases (MMPs) are capable of degrading extracellular matrix, and they are inducible enzymes depending on an inflammatory environment such as periodontitis and bacterial infection in periodontal tissue. Gingival inflammation has been postulated to be correlated with the production of MMP-2 and MMP-9. The objective of this study was to quantify the expression and activity of MMP-9 and -2, and to determine the correlation between activity and expression of these MMPs in human gingival tissues with periodontitis. The gingival tissues of 13 patients were homogenized in 500 µL of phosphate buffered saline with a protease inhibitor cocktail. The expression and activity of MMP-2 and -9 were measured by enzyme-linked immunosorbent assay and Western blot analysis, and quantified by a densitometer. For the correlation line, statistical analysis was performed using the Systat software package. MMP-9 was highly expressed in all gingival tissue samples, whereas MMP-2 was underexpressed compared with MMP-9. MMP-9 activity increased together with the MMP-9 expression level, with a positive correlation (r=0.793, P=0.01). The correlation was not observed in MMP-2. The expression of MMP-2 and -9 might contribute to periodontal physiological and pathological processes, and the degree of MMP-9 expression and activity are predictive indicators relevant to the progression of periodontitis.

  13. Rodent neonatal germinal matrix hemorrhage mimics the human brain injury, neurological consequences, and post-hemorrhagic hydrocephalus.

    Science.gov (United States)

    Lekic, Tim; Manaenko, Anatol; Rolland, William; Krafft, Paul R; Peters, Regina; Hartman, Richard E; Altay, Orhan; Tang, Jiping; Zhang, John H

    2012-07-01

    Germinal matrix hemorrhage (GMH) is the most common neurological disease of premature newborns. GMH causes neurological sequelae such as cerebral palsy, post-hemorrhagic hydrocephalus, and mental retardation. Despite this, there is no standardized animal model of spontaneous GMH using newborn rats to depict the condition. We asked whether stereotactic injection of collagenase type VII (0.3 U) into the ganglionic eminence of neonatal rats would reproduce the acute brain injury, gliosis, hydrocephalus, periventricular leukomalacia, and attendant neurological consequences found in humans. To test this hypothesis, we used our neonatal rat model of collagenase-induced GMH in P7 pups, and found that the levels of free-radical adducts (nitrotyrosine and 4-hyroxynonenal), proliferation (mammalian target of rapamycin), inflammation (COX-2), blood components (hemoglobin and thrombin), and gliosis (vitronectin and GFAP) were higher in the forebrain of GMH pups, than in controls. Neurobehavioral testing showed that pups with GMH had developmental delay, and the juvenile animals had significant cognitive and motor disability, suggesting clinical relevance of the model. There was also evidence of white-matter reduction, ventricular dilation, and brain atrophy in the GMH animals. This study highlights an instructive animal model of the neurological consequences after germinal matrix hemorrhage, with evidence of brain injuries that can be used to evaluate strategies in the prevention and treatment of post-hemorrhagic complications. Copyright © 2012 Elsevier Inc. All rights reserved.

  14. In vitro comparison of new bisphosphonic acids and zoledronate effects on human gingival fibroblasts viability, inflammation and matrix turnover.

    Science.gov (United States)

    De Colli, Marianna; Tortorella, Paolo; Marconi, Guya Diletta; Agamennone, Mariangela; Campestre, Cristina; Tauro, Marilena; Cataldi, Amelia; Zara, Susi

    2016-11-01

    Bisphosphonates (BPs) are drugs clinically used in resorptive diseases. It was already proved that some clinically relevant BPs can inhibit a class of enzymes called matrix metalloproteinases (MMPs), required during tissue remodelling. Combining the arylsulfonamide function with the bisphosphonic group, several compounds were synthesized to obtain selective inhibitors of MMPs. The aim of the present study was to compare the effect of zoledronic acid (ZA), the most potent bisphosphonate available as therapy, with new sulfonamide containing BPs in an in vitro model of human gingival fibroblasts (HGFs). Western blot was used to measure procollagen I, β1 integrin MMP-8 and MMP-9, phase contrast and MTT for cell viability; L-lactate-dehydrogenase (LDH) measurement was performed for toxicity evaluation and ELISA for prostaglandin E 2 (PGE 2 ) secretion assessment. When compared with ZA, the treatment with the newly synthesized compounds shows increasing viability, procollagen I expression and decreased expression of β1 integrin in HGFs. Higher levels of released LDH, PGE 2 and MMP-9 expression are recorded in ZA-treated HGFs. Increased levels of MMP-8 are recorded in newly synthesized compounds-treated samples. These findings allowed to conclude that new tested BPs did not affect HGFs viability and adhesion, did not induce cellular toxicity, were not responsible for inflammatory event induction and could preserve the physiological matrix turnover. It could be hypothesized that the new molecules were better tolerated by soft tissues, resulting in lesser side effects.

  15. Skip Regulates TGF-β1-Induced Extracellular Matrix Degrading Proteases Expression in Human PC-3 Prostate Cancer Cells

    Directory of Open Access Journals (Sweden)

    Victor Villar

    2013-01-01

    Full Text Available Purpose. To determine whether Ski-interacting protein (SKIP regulates TGF-β1-stimulated expression of urokinase-type plasminogen activator (uPA, matrix metalloproteinase-9 (MMP-9, and uPA Inhibitor (PAI-1 in the androgen-independent human prostate cancer cell model. Materials and Methods. PC-3 prostate cancer cell line was used. The role of SKIP was evaluated using synthetic small interference RNA (siRNA compounds. The expression of uPA, MMP-9, and PAI-1 was evaluated by zymography assays, RT-PCR, and promoter transactivation analysis. Results. In PC-3 cells TGF-β1 treatment stimulated uPA, PAI-1, and MMP-9 expressions. The knockdown of SKIP in PC-3 cells enhanced the basal level of uPA, and TGF-β1 treatment inhibited uPA production. Both PAI-1 and MMP-9 production levels were increased in response to TGF-β1. The ectopic expression of SKIP inhibited both TGF-β1-induced uPA and MMP-9 promoter transactivation, while PAI-1 promoter response to the factor was unaffected. Conclusions. SKIP regulates the expression of uPA, PAI-1, and MMP-9 stimulated by TGF-β1 in PC-3 cells. Thus, SKIP is implicated in the regulation of extracellular matrix degradation and can therefore be suggested as a novel therapeutic target in prostate cancer treatment.

  16. The impact of the matrix of red beet products and interindividual variability on betacyanins bioavailability in humans.

    Science.gov (United States)

    Wiczkowski, Wieslaw; Romaszko, Ewa; Szawara-Nowak, Dorota; Piskula, Mariusz K

    2018-06-01

    The influence of the matrix of red beetroot products and interindividual variability on betacyanins bioavailability in humans was studied. In a randomized crossover study 12 volunteers consumed red beet juice and crunchy slices containing betanin and isobetanin. Betalains were analyzed by the HPLC-DAD-MS. Urine samples examined after the consumption of both products contained not only native betacyanins but also their aglycones. In case of juice, the highest betacyanins urine excretion rate was observed within the first 2 h (64 nmol/h), while in case of crunchy slices within the period of 2-4 h (66 nmol/h). Among volunteers, the average total betacyanins excretion rate ranged from 18.54 to 67.96 nmol/h and, 13.15 to 63.58 nmol/h for red beet juice and crunchy slices, respectively. In total, approximately 0.3% of betacyanins (ranging from 0.12 to 0.58%) ingested from both products was excreted. The study showed that betacyanins bioavailability from juice and crunchy slices is similar, with the matrix of products consumed having an impact on betacyanins excretion profile, and the phenotype of volunteers affecting betacyanins excretion rate. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. Human alveolar bone cell proliferation, expression of osteoblastic phenotype, and matrix mineralization on porous titanium produced by powder metallurgy.

    Science.gov (United States)

    Rosa, Adalberto Luiz; Crippa, Grasiele Edilaine; de Oliveira, Paulo Tambasco; Taba, Mario; Lefebvre, Louis-Philippe; Beloti, Marcio Mateus

    2009-05-01

    This study aimed at investigating the influence of the porous titanium (Ti) structure on the osteogenic cell behaviour. Porous Ti discs were fabricated by the powder metallurgy process with the pore size typically between 50 and 400 microm and a porosity of 60%. Osteogenic cells obtained from human alveolar bone were cultured until subconfluence and subcultured on dense Ti (control) and porous Ti for periods of up to 17 days. Cultures grown on porous Ti exhibited increased cell proliferation and total protein content, and lower levels of alkaline phosphatase (ALP) activity than on dense Ti. In general, gene expression of osteoblastic markers-runt-related transcription factor 2, collagen type I, alkaline phosphatase, bone morphogenetic protein-7, and osteocalcin was lower at day 7 and higher at day 17 in cultures grown on porous Ti compared with dense Ti, a finding consistent with the enhanced growth rate for such cultures. The amount of mineralized matrix was greater on porous Ti compared with the dense one. These results indicate that the porous Ti is an appropriate substrate for osteogenic cell adhesion, proliferation, and production of a mineralized matrix. Because of the three-dimensional environment it provides, porous Ti should be considered an advantageous substrate for promoting desirable implant surface-bone interactions.

  18. Tantalum coating on TiO{sub 2} nanotubes induces superior rate of matrix mineralization and osteofunctionality in human osteoblasts

    Energy Technology Data Exchange (ETDEWEB)

    Frandsen, Christine J.; Brammer, Karla S. [Materials Science and Engineering, University of California at San Diego, La Jolla, CA 92093 (United States); Noh, Kunbae [Corporate Research Institute, Cheil Industries, Inc., Gocheon-Dong, Uiwang-Si, Gyeonggi-Do, 437-711 (Korea, Republic of); Johnston, Gary [Materials Science and Engineering, University of California at San Diego, La Jolla, CA 92093 (United States); Jin, Sungho, E-mail: jin@ucsd.edu [Materials Science and Engineering, University of California at San Diego, La Jolla, CA 92093 (United States); Mechanical and Aerospace Engineering, University of California at San Diego, La Jolla, CA 92093 (United States)

    2014-04-01

    Nanostructured surface geometries have been the focus of a multitude of recent biomaterial research, and exciting findings have been published. However, only a few publications have directly compared nanostructures of various surface chemistries. The work herein directly compares the response of human osteoblast cells to surfaces of identical nanotube geometries with two well-known orthopedic biomaterials: titanium oxide (TiO{sub 2}) and tantalum (Ta). The results reveal that the Ta surface chemistry on the nanotube architecture enhances alkaline phosphatase activity, and promotes a ∼ 30% faster rate of matrix mineralization and bone-nodule formation when compared to results on bare TiO{sub 2} nanotubes. This study implies that unique combinations of surface chemistry and nanostructure may influence cell behavior due to distinctive physico-chemical properties. These findings are of paramount importance to the orthopedics field for understanding cell behavior in response to subtle alterations in nanostructure and surface chemistry, and will enable further insight into the complex manipulation of biomaterial surfaces. With increased focus in the field of orthopedic materials research on nanostructured surfaces, this study emphasizes the need for careful and systematic review of variations in surface chemistry in concurrence with nanotopographical changes. - Highlights: • A TiO{sub 2} nanotube surface structure was coated with tantalum. • Osteoblast cell response was compared between the tantalum coated and as-formed TiO{sub 2} nanotube surface. • We observed superior rates of bone matrix mineralization and osteoblast maturation on the tantalum coated nanotube surface.

  19. Tantalum coating on TiO2 nanotubes induces superior rate of matrix mineralization and osteofunctionality in human osteoblasts

    International Nuclear Information System (INIS)

    Frandsen, Christine J.; Brammer, Karla S.; Noh, Kunbae; Johnston, Gary; Jin, Sungho

    2014-01-01

    Nanostructured surface geometries have been the focus of a multitude of recent biomaterial research, and exciting findings have been published. However, only a few publications have directly compared nanostructures of various surface chemistries. The work herein directly compares the response of human osteoblast cells to surfaces of identical nanotube geometries with two well-known orthopedic biomaterials: titanium oxide (TiO 2 ) and tantalum (Ta). The results reveal that the Ta surface chemistry on the nanotube architecture enhances alkaline phosphatase activity, and promotes a ∼ 30% faster rate of matrix mineralization and bone-nodule formation when compared to results on bare TiO 2 nanotubes. This study implies that unique combinations of surface chemistry and nanostructure may influence cell behavior due to distinctive physico-chemical properties. These findings are of paramount importance to the orthopedics field for understanding cell behavior in response to subtle alterations in nanostructure and surface chemistry, and will enable further insight into the complex manipulation of biomaterial surfaces. With increased focus in the field of orthopedic materials research on nanostructured surfaces, this study emphasizes the need for careful and systematic review of variations in surface chemistry in concurrence with nanotopographical changes. - Highlights: • A TiO 2 nanotube surface structure was coated with tantalum. • Osteoblast cell response was compared between the tantalum coated and as-formed TiO 2 nanotube surface. • We observed superior rates of bone matrix mineralization and osteoblast maturation on the tantalum coated nanotube surface

  20. Quantitative proteomics reveals altered expression of extracellular matrix related proteins of human primary dermal fibroblasts in response to sulfated hyaluronan and collagen applied as artificial extracellular matrix.

    Science.gov (United States)

    Müller, Stephan A; van der Smissen, Anja; von Feilitzsch, Margarete; Anderegg, Ulf; Kalkhof, Stefan; von Bergen, Martin

    2012-12-01

    Fibroblasts are the main matrix producing cells of the dermis and are also strongly regulated by their matrix environment which can be used to improve and guide skin wound healing processes. Here, we systematically investigated the molecular effects on primary dermal fibroblasts in response to high-sulfated hyaluronan [HA] (hsHA) by quantitative proteomics. The comparison of non- and high-sulfated HA revealed regulation of 84 of more than 1,200 quantified proteins. Based on gene enrichment we found that sulfation of HA alters extracellular matrix remodeling. The collagen degrading enzymes cathepsin K, matrix metalloproteinases-2 and -14 were found to be down-regulated on hsHA. Additionally protein expression of thrombospondin-1, decorin, collagen types I and XII were reduced, whereas the expression of trophoblast glycoprotein and collagen type VI were slightly increased. This study demonstrates that global proteomics provides a valuable tool for revealing proteins involved in molecular effects of growth substrates for further material optimization.

  1. A model for cell migration in non-isotropic fibrin networks with an application to pancreatic tumor islets.

    Science.gov (United States)

    Chen, Jiao; Weihs, Daphne; Vermolen, Fred J

    2018-04-01

    Cell migration, known as an orchestrated movement of cells, is crucially important for wound healing, tumor growth, immune response as well as other biomedical processes. This paper presents a cell-based model to describe cell migration in non-isotropic fibrin networks around pancreatic tumor islets. This migration is determined by the mechanical strain energy density as well as cytokines-driven chemotaxis. Cell displacement is modeled by solving a large system of ordinary stochastic differential equations where the stochastic parts result from random walk. The stochastic differential equations are solved by the use of the classical Euler-Maruyama method. In this paper, the influence of anisotropic stromal extracellular matrix in pancreatic tumor islets on T-lymphocytes migration in different immune systems is investigated. As a result, tumor peripheral stromal extracellular matrix impedes the immune response of T-lymphocytes through changing direction of their migration.

  2. Extracellular Matrix Modulates Morphology, Growth, Oxidative Stress Response and Functionality of Human Skin Fibroblasts during Aging In Vitro

    DEFF Research Database (Denmark)

    Jørgensen, Peter; Rattan, Suresh

    2014-01-01

    recent observations indicate that replicative lifespan, senescence and functionality of cells in vitro can be significantly affected by the quality of the extra cellular matrix (ECM). Following up on those reports, here we show that using the ECM prepared from early passage young cells, partial...... rejuvenation of serially passaged human facial skin fibroblasts was possible in pre-senescent middle-aged cells, but not in fully senescent late passage cells. ECM from young cells improved the appearance, viability, stress tolerance and wound healing ability of skin fibroblasts. Furthermore, young ECM...... modulated the oxidative stress response transcription factor Nrf-2 and its downstream effector haem-oxygenase (HO-1), possibly through the amelioration of the environmental stress induced by the plastic surface of the culturing flasks. Therefore, it is important to consider the role of ECM in modulating...

  3. The tissue-engineered human cornea as a model to study expression of matrix metalloproteinases during corneal wound healing.

    Science.gov (United States)

    Couture, Camille; Zaniolo, Karine; Carrier, Patrick; Lake, Jennifer; Patenaude, Julien; Germain, Lucie; Guérin, Sylvain L

    2016-02-01

    Corneal injuries remain a major cause of consultation in the ophthalmology clinics worldwide. Repair of corneal wounds is a complex mechanism that involves cell death, migration, proliferation, differentiation, and extracellular matrix (ECM) remodeling. In the present study, we used a tissue-engineered, two-layers (epithelium and stroma) human cornea as a biomaterial to study both the cellular and molecular mechanisms of wound healing. Gene profiling on microarrays revealed important alterations in the pattern of genes expressed by tissue-engineered corneas in response to wound healing. Expression of many MMPs-encoding genes was shown by microarray and qPCR analyses to increase in the migrating epithelium of wounded corneas. Many of these enzymes were converted into their enzymatically active form as wound closure proceeded. In addition, expression of MMPs by human corneal epithelial cells (HCECs) was affected both by the stromal fibroblasts and the collagen-enriched ECM they produce. Most of all, results from mass spectrometry analyses provided evidence that a fully stratified epithelium is required for proper synthesis and organization of the ECM on which the epithelial cells adhere. In conclusion, and because of the many characteristics it shares with the native cornea, this human two layers corneal substitute may prove particularly useful to decipher the mechanistic details of corneal wound healing. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Generation of biologically active endostatin fragments from human collagen XVIII by distinct matrix metalloproteases

    International Nuclear Information System (INIS)

    Heljasvaara, Ritva; Nyberg, Pia; Luostarinen, Jani; Parikka, Mataleena; Heikkilae, Pia; Rehn, Marko; Sorsa, Timo; Salo, Tuula; Pihlajaniemi, Taina

    2005-01-01

    Endostatin, a potent inhibitor of endothelial cell proliferation, migration, angiogenesis and tumor growth, is proteolytically cleaved from the C-terminal noncollagenous NC1 domain of type XVIII collagen. We investigated the endostatin formation from human collagen XVIII by several MMPs in vitro. The generation of endostatin fragments differing in molecular size (24-30 kDa) and in N-terminal sequences was identified in the cases of MMP-3, -7, -9, -13 and -20. The cleavage sites were located in the protease-sensitive hinge region between the trimerization and endostatin domains of NC1. MMP-1, -2, -8 and -12 did not show any significant activity against the C-terminus of collagen XVIII. The anti-proliferative effect of the 20-kDa endostatin, three longer endostatin-containing fragments generated in vitro by distinct MMPs and the entire NC1 domain, on bFGF-stimulated human umbilical vein endothelial cells was established. The anti-migratory potential of some of these fragments was also studied. In addition, production of endostatin fragments between 24-30 kDa by human hepatoblastoma cells was shown to be due to MMP action on type XVIII collagen. Our results indicate that certain, especially cancer-related, MMP family members can generate biologically active endostatin-containing polypeptides from collagen XVIII and thus, by releasing endostatin fragments, may participate in the inhibition of endothelial cell proliferation, migration and angiogenesis

  5. [Eosinophilic pleural effusion possibly induced by fibrin sealant].

    Science.gov (United States)

    Kambayashi, Takatoyo; Suzuki, Takashi

    2012-02-01

    A 74-year-old man underwent right upper lobectomy for the lung cancer and bullectomy of right lower lobe. Fibrin sealant was used for sealing the excision line. The increase of the pleural effusion with increasing C-reactive protein( CRP) and eosinophilia was noted at the 17th day after the operation. The pleural effusion was transparent and yellowish colored suggesting transudatory liquid. The eosinophil in the pleural effusion was as high as 14%. The drainage of the pleural effusion was performed for 2 days resulting in disappearing the abnormal accumulation without any additional treatment. The cause of pleural effusion was supposed to be fibrin sealant by a positive result of the drug lymphocyte stimulation test.

  6. A novel functional site of extracellular matrix metalloproteinase inducer (EMMPRIN) that limits the migration of human uterine cervical carcinoma cells.

    Science.gov (United States)

    Sato, Takashi; Watanabe, Mami; Hashimoto, Kei; Ota, Tomoko; Akimoto, Noriko; Imada, Keisuke; Nomizu, Motoyoshi; Ito, Akira

    2012-01-01

    EMMPRIN (extracellular matrix metalloproteinase inducer)/CD147, a membrane-bound glycoprotein with two extracellular loop domains (termed loops I and II), progresses tumor invasion and metastasis by increasing the production of matrix metalloproteinase (MMP) in peritumoral stoma cells. EMMPRIN has also been associated with the control of migration activity in some tumor cells, but little is known about how EMMPRIN regulates tumor cell migration. In the present study, EMMPRIN siRNA suppressed the gene expression and production of EMMPRIN in human uterine cervical carcinoma SKG-II cells. An in vitro scratch wound assay showed enhancement of migration of EMMPRIN-knockdown SKG-II cells. In addition, the SKG-II cell migration was augmented by adding an E. coli-expressed human EMMPRIN mutant with two extracellular loop domains (eEMP-I/II), which bound to the cell surface of SKG-II cells. However, eEMP-I/II suppressed the native EMMPRIN-mediated augmentation of proMMP-1/procollagenase-1 production in a co-culture of the SKG-II cells and human uterine cervical fibroblasts, indicating that the augmentation of SKG-II cell migration resulted from the interference of native EMMPRIN functions by eEMP-I/II on the cell surface. Furthermore, a systematic peptide screening method using nine synthetic EMMPRIN peptides coding the loop I and II domains (termed EM1-9) revealed that EM9 (170HIENLNMEADPGQYR184) facilitated SKG-II cell migration. Moreover, SKG-II cell migration was enhanced by administration of an antibody against EM9, but not EM1 which is a crucial site for the MMP inducible activity of EMMPRIN. Therefore, these results provide novel evidence that EMMPRIN on the cell surface limits the cell migration of human uterine cervical carcinoma cells through 170HIENLNMEADPGQYR184 in the loop II domain. Finally, these results should provide an increased understanding of the functions of EMMPRIN in malignant cervical carcinoma cells, and could contribute to the development of

  7. Two Distinct Isoforms of Matrix Metalloproteinase-2 Are Associated with Human Delayed Kidney Graft Function.

    Directory of Open Access Journals (Sweden)

    Shaynah Wanga

    Full Text Available Delayed graft function (DGF is a frequent complication of renal transplantation, particularly in the setting of transplantation of kidneys derived from deceased donors and expanded-criteria donors. DGF results from tubular epithelial cell injury and has immediate and long term consequences. These include requirement for post-transplantation dialysis, increased incidence of acute rejection, and poorer long-term outcomes. DGF represents one of the clearest clinical examples of renal acute ischemia/reperfusion injury. Experimental studies have demonstrated that ischemia/reperfusion injury induces the synthesis of the full length secreted isoform of matrix metalloproteinase-2 (FL-MMP-2, as well as an intracellular N-terminal truncated MMP-2 isoform (NTT-MMP-2 that initiates an innate immune response. We hypothesized that the two MMP-2 isoforms mediate tubular epithelial cell injury in DGF. Archival renal biopsy sections from 10 protocol biopsy controls and 41 cases with a clinical diagnosis of DGF were analyzed for the extent of tubular injury, expression of the FL-MMP-2 and NTT-MMP-2 isoforms by immunohistochemistry (IHC, in situ hybridization, and qPCR to determine isoform abundance. Differences in transcript abundance were related to tubular injury score. Markers of MMP-2-mediated injury included TUNEL staining and assessment of peritubular capillary density. There was a clear relationship between tubular epithelial cell expression of both FL-MMP-2 and NTT-MMP-2 IHC with the extent of tubular injury. The MMP-2 isoforms were detected in the same tubular segments and were present at sites of tubular injury. qPCR demonstrated highly significant increases in both the FL-MMP-2 and NTT-MMP-2 transcripts. Statistical analysis revealed highly significant associations between FL-MMP-2 and NTT-MMP-2 transcript abundance and the extent of tubular injury, with NTT-MMP-2 having the strongest association. We conclude that two distinct MMP-2 isoforms are

  8. Physical determinants of fibrinolysis in single fibrin fibers.

    Directory of Open Access Journals (Sweden)

    Igal Bucay

    Full Text Available Fibrin fibers form the structural backbone of blood clots; fibrinolysis is the process in which plasmin digests fibrin fibers, effectively regulating the size and duration of a clot. To understand blood clot dissolution, the influence of clot structure and fiber properties must be separated from the effects of enzyme kinetics and perfusion rates into clots. Using an inverted optical microscope and fluorescently-labeled fibers suspended between micropatterned ridges, we have directly measured the lysis of individual fibrin fibers. We found that during lysis 64 ± 6% of fibers were transected at one point, but 29 ± 3% of fibers increase in length rather than dissolving or being transected. Thrombin and plasmin dose-response experiments showed that the elongation behavior was independent of plasmin concentration, but was instead dependent on the concentration of thrombin used during fiber polymerization, which correlated inversely with fiber diameter. Thinner fibers were more likely to lyse, while fibers greater than 200 ± 30 nm in diameter were more likely to elongate. Because lysis rates were greatly reduced in elongated fibers, we hypothesize that plasmin activity depends on fiber strain. Using polymer physics- and continuum mechanics-based mathematical models, we show that fibers polymerize in a strained state and that thicker fibers lose their prestrain more rapidly than thinner fibers during lysis, which may explain why thick fibers elongate and thin fibers lyse. These results highlight how subtle differences in the diameter and prestrain of fibers could lead to dramatically different lytic susceptibilities.

  9. Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

    International Nuclear Information System (INIS)

    Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki; Kondo, Ayami; Nakata, Kazuhiko; Mogi, Makio

    2015-01-01

    A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CM treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration

  10. AMP-activated protein kinase activation mediates CCL3-induced cell migration and matrix metalloproteinase-2 expression in human chondrosarcoma

    Science.gov (United States)

    2013-01-01

    Chemokine (C-C motif) ligand 3 (CCL3), also known as macrophage inflammatory protein-1α, is a cytokine involved in inflammation and activation of polymorphonuclear leukocytes. CCL3 has been detected in infiltrating cells and tumor cells. Chondrosarcoma is a highly malignant tumor that causes distant metastasis. However, the effect of CCL3 on human chondrosarcoma metastasis is still unknown. Here, we found that CCL3 increased cellular migration and expression of matrix metalloproteinase (MMP)-2 in human chondrosarcoma cells. Pre-treatment of cells with the MMP-2 inhibitor or transfection with MMP-2 specific siRNA abolished CCL3-induced cell migration. CCL3 has been reported to exert its effects through activation of its specific receptor, CC chemokine receptor 5 (CCR5). The CCR5 and AMP-activated protein kinase (AMPK) inhibitor or siRNA also attenuated CCL3-upregulated cell motility and MMP-2 expression. CCL3-induced expression of MMP-2 and migration were also inhibited by specific inhibitors, and inactive mutants of AMPK, p38 mitogen activated protein kinase (p38 or p38-MAPK), and nuclear factor κB (NF-κB) cascades. On the other hand, CCL3 treatment demonstrably activated AMPK, p38, and NF-κB signaling pathways. Furthermore, the expression levels of CCL3, CCR5, and MMP-2 were correlated in human chondrosarcoma specimens. Taken together, our results indicate that CCL3 enhances the migratory ability of human chondrosarcoma cells by increasing MMP-2 expression via the CCR5, AMPK, p38, and NF-κB pathways. PMID:24047437

  11. Development of matrix solid-phase dispersion method for the extraction of short-chain chlorinated paraffins in human placenta.

    Science.gov (United States)

    Wang, Ying; Gao, Wei; Wu, Jing; Liu, Huijin; Wang, Yingjun; Wang, Yawei; Jiang, Guibin

    2017-12-01

    Chlorinated paraffins (SCCPs) are widely used worldwide, and they can be released into the environment during their production, transport, usage and disposal, which pose potential risks for human health. In this work, an efficient, reliable and rapid pretreatment method based on matrix solid-phase dispersion (MSPD) was developed for the analysis of short-chain CPs (SCCPs) in human placenta by gas chromatograph-electron capture negative ion low-resolution mass spectrometry (GC-ECNI-LRMS) and gas chromatography-quadrupole time-of-flight mass spectrometry (GC-QTOF-HRMS). The MSPD-relevant parameters including dispersing sorbent, sample-to-sorbent mass ratio, and elution solvent were optimized using the orthogonal test. Silica gel was found to be the optimal dispersing sorbent among the selected matrices. Under the optimal conditions, 44% acidic silica gel can be used as the co-sorbent to remove lipid and eluted by the mixture of hexane and dichloromethane (7:3, V/V). The spiked recoveries of the optimized method were 77.4% and 91.4% for analyzing SCCPs in human placenta by GC-ECNI-LRMS and GC-QTOF-HRMS, and the corresponding relative standard deviations were 10.2% and 5.6%, respectively. The method detection limit for the total SCCPs was 36.8ng/g (dry weight, dw) and 19.2ng/g (dw) as measured by GC-ECNI-LRMS and GC-QTOF-HRMS, respectively. The concentrations of SCCPs in four human placentas were in the range of

  12. Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

    Energy Technology Data Exchange (ETDEWEB)

    Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya 464-8651, Aichi (Japan); Kondo, Ayami [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan); Nakata, Kazuhiko [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya 464-8651, Aichi (Japan); Mogi, Makio, E-mail: makio@dpc.agu.ac.jp [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan)

    2015-02-01

    A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CM treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration.

  13. Thiamine and benfotiamine prevent apoptosis induced by high glucose-conditioned extracellular matrix in human retinal pericytes.

    Science.gov (United States)

    Beltramo, Elena; Nizheradze, Konstantin; Berrone, Elena; Tarallo, Sonia; Porta, Massimo

    2009-10-01

    Early and selective loss of pericytes and thickening of the basement membrane are hallmarks of diabetic retinopathy. We reported reduced adhesion, but no changes in apoptosis, of bovine retinal pericytes cultured on extracellular matrix (ECM) produced by endothelial cells in high glucose (HG). Since human and bovine pericytes may behave differently in conditions mimicking the diabetic milieu, we verified the behaviour of human retinal pericytes cultured on HG-conditioned ECM. Pericytes were cultured in physiological/HG on ECM produced by human umbilical vein endothelial cells in physiological/HG, alone or in the presence of thiamine and benfotiamine. Adhesion, proliferation, apoptosis, p53 and Bcl-2/Bax ratio (mRNA levels and protein concentrations) were measured in wild-type and immortalized human pericytes. Both types of pericytes adhered less to HG-conditioned ECM and plastic than to physiological glucose-conditioned ECM. DNA synthesis was impaired in pericytes cultured in HG on the three different surfaces but there were no differences in proliferation. DNA fragmentation and Bcl-2/Bax ratio were greatly enhanced by HG-conditioned ECM in pericytes kept in both physiological and HG. Addition of thiamine and benfotiamine to HG during ECM production completely prevented these damaging effects. Apoptosis is strongly increased in pericytes cultured on ECM produced by endothelium in HG, probably due to impairment of the Bcl-2/Bax ratio. Thiamine and benfotiamine completely revert this effect. This behaviour is therefore completely different from that of bovine pericytes, underlining the importance of establishing species-specific cell models to study the mechanisms of diabetic retinopathy. (c) 2009 John Wiley & Sons, Ltd.

  14. The action characterization matrix: A link between HERA (Human Events Reference for ATHEANA) and ATHEANA (a technique for human error analysis)

    International Nuclear Information System (INIS)

    Hahn, H.A.

    1997-01-01

    The Technique for Human Error Analysis (ATHEANA) is a newly developed human reliability analysis (HRA) methodology that aims to facilitate better representation and integration of human performance into probabilistic risk assessment (PRA) modeling and quantification by analyzing risk-significant operating experience in the context of existing behavior science models. The fundamental premise of ATHEANA is that error-forcing contexts (EFCs), which refer to combinations of equipment/material conditions and performance shaping factors (PSFs), set up or create the conditions under which unsafe actions (UAs) can occur. ATHEANA is being developed in the context of nuclear power plant (NPP) PRAs, and much of the language used to describe the method and provide examples of its application are specific to that industry. Because ATHEANA relies heavily on the analysis of operational events that have already occurred as a mechanism for generating creative thinking about possible EFCs, a database, called the Human Events Reference for ATHEANA (HERA), has been developed to support the methodology. Los Alamos National Laboratory's (LANL) Human Factors Group has recently joined the ATHEANA project team; LANL is responsible for further developing the database structure and for analyzing additional exemplar operational events for entry into the database. The Action Characterization Matrix (ACM) is conceived as a bridge between the HERA database structure and ATHEANA. Specifically, the ACM allows each unsafe action or human failure event to be characterized according to its representation along each of six different dimensions: system status, initiator status, unsafe action mechanism, information processing stage, equipment/material conditions, and performance shaping factors. This report describes the development of the ACM and provides details on the structure and content of its dimensions

  15. Enhanced transfection efficiency of human embryonic stem cells by the incorporation of DNA liposomes in extracellular matrix.

    Science.gov (United States)

    Villa-Diaz, Luis G; Garcia-Perez, Jose L; Krebsbach, Paul H

    2010-12-01

    Because human embryonic stem (hES) cells can differentiate into virtually any cell type in the human body, these cells hold promise for regenerative medicine. The genetic manipulation of hES cells will enhance our understanding of genes involved in early development and will accelerate their potential use and application for regenerative medicine. The objective of this study was to increase the transfection efficiency of plasmid DNA into hES cells by modifying a standard reverse transfection (RT) protocol of lipofection. We hypothesized that immobilization of plasmid DNA in extracellular matrix would be a more efficient method for plasmid transfer due to the affinity of hES cells for substrates such as Matrigel and to the prolonged exposure of cells to plasmid DNA. Our results demonstrate that this modification doubled the transfection efficiency of hES cells and the generation of clonal cell lines containing a piece of foreign DNA stably inserted in their genomes compared to results obtained with standard forward transfection. In addition, treatment with dimethyl sulfoxide further increased the transfection efficiency of hES cells. In conclusion, modifications to the RT protocol of lipofection result in a significant and robust increase in the transfection efficiency of hES cells.

  16. Inhibiting Invasion into Human Bladder Carcinoma 5637 Cells with Diallyl Trisulfide by Inhibiting Matrix Metalloproteinase Activities and Tightening Tight Junctions

    Directory of Open Access Journals (Sweden)

    Yung Hyun Choi

    2013-10-01

    Full Text Available Diallyl trisulfide (DATS, an organosulfur compound in garlic, possesses pronounced anti-cancer potential. However, the anti-invasive mechanism of this compound in human bladder carcinoma is not fully understood. In this study, we evaluated the anti-invasive effects of DATS on a human bladder carcinoma (5637 cell line and investigated the underlying mechanism. The results indicated that DATS suppressed migration and invasion of 5637 cells by reducing the activities and expression of matrix metalloproteinase (MMP-2 and MMP-9 at both the protein and mRNA levels. DATS treatment up-regulated expression of tissue inhibitor of metalloproteinase (TIMP-1 and TIMP-2 in 5637 cells. The inhibitory effects of DATS on invasiveness were associated with an increase in transepithelial electrical resistance and repression of the levels of claudin family members. Although further studies are needed, our data demonstrate that DATS exhibits anti-invasive effects in 5637 cells by down-regulating the activity of tight junctions and MMPs. DATS may have future utility in clinical applications for treating bladder cancer.

  17. Bioprinting 3D cell-laden hydrogel microarray for screening human periodontal ligament stem cell response to extracellular matrix

    International Nuclear Information System (INIS)

    Ma, Yufei; Ji, Yuan; Huang, Guoyou; Zhang, Xiaohui; Xu, Feng; Ling, Kai

    2015-01-01

    Periodontitis is an inflammatory disease negatively affecting up to 15% of adults worldwide. Periodontal ligament stem cells (PDLSCs) hold great promises for periodontal tissue regeneration, where it is necessary to find proper extracellular matrix (ECM) materials (e.g., composition, concentration). In this study, we proposed a bioprinting-based approach to generate nano-liter sized three-dimensional (3D) cell-laden hydrogel array with gradient of ECM components, through controlling the volume ratio of two hydrogels, such as gelatin methacrylate (GelMA) and poly(ethylene glycol) (PEG) dimethacrylate. The resulting cell-laden array with a gradient of GelMA/PEG composition was used to screen human PDLSC response to ECM. The behavior (e.g., cell viability, spreading) of human PDLSCs in GelMA/PEG array were found to be depended on the volume ratios of GelMA/PEG, with cell viability and spreading area decreased along with increasing the ratio of PEG. The developed approach would be useful for screening cell-biomaterial interaction in 3D and promoting regeneration of functional tissue. (paper)

  18. Regeneration of Cartilage in Human Knee Osteoarthritis with Autologous Adipose Tissue-Derived Stem Cells and Autologous Extracellular Matrix

    Directory of Open Access Journals (Sweden)

    Jaewoo Pak

    2016-08-01

    Full Text Available This clinical case series demonstrates that percutaneous injections of autologous adipose tissue-derived stem cells (ADSCs and homogenized extracellular matrix (ECM in the form of adipose stromal vascular fraction (SVF, along with hyaluronic acid (HA and platelet-rich plasma (PRP activated by calcium chloride, could regenerate cartilage-like tissue in human knee osteoarthritis (OA patients. Autologous lipoaspirates were obtained from adipose tissue of the abdominal origin. Afterward, the lipoaspirates were minced to homogenize the ECM. These homogenized lipoaspirates were then mixed with collagenase and incubated. The resulting mixture of ADSCs and ECM in the form of SVF was injected, along with HA and PRP activated by calcium chloride, into knees of three Korean patients with OA. The same affected knees were reinjected weekly with additional PRP activated by calcium chloride for 3 weeks. Pretreatment and post-treatment magnetic resonance imaging (MRI data, functional rating index, range of motion (ROM, and pain score data were then analyzed. All patients' MRI data showed cartilage-like tissue regeneration. Along with MRI evidence, the measured physical therapy outcomes in terms of ROM, subjective pain, and functional status were all improved. This study demonstrates that percutaneous injection of ADSCs with ECM contained in autologous adipose SVF, in conjunction with HA and PRP activated by calcium chloride, is a safe and potentially effective minimally invasive therapy for OA of human knees.

  19. Proteomic Analysis of Human Tendon and Ligament: Solubilization and Analysis of Insoluble Extracellular Matrix in Connective Tissues.

    Science.gov (United States)

    Sato, Nori; Taniguchi, Takako; Goda, Yuichiro; Kosaka, Hirofumi; Higashino, Kosaku; Sakai, Toshinori; Katoh, Shinsuke; Yasui, Natsuo; Sairyo, Koichi; Taniguchi, Hisaaki

    2016-12-02

    Connective tissues such as tendon, ligament and cartilage are mostly composed of extracellular matrix (ECM). These tissues are insoluble, mainly due to the highly cross-linked ECM proteins such as collagens. Difficulties obtaining suitable samples for mass spectrometric analysis render the application of modern proteomic technologies difficult. Complete solubilization of them would not only elucidate protein composition of normal tissues but also reveal pathophysiology of pathological tissues. Here we report complete solubilization of human Achilles tendon and yellow ligament, which is achieved by chemical digestion combined with successive protease treatment including elastase. The digestion mixture was subjected to liquid chromatography-mass spectrometry. The low specificity of elastase was overcome by accurate mass analysis achieved using FT-ICR-MS. In addition to the detailed proteome of both tissues, we also quantitatively determine the major protein composition of samples, by measuring peak area of some characteristic peptides detected in tissue samples and in purified proteins. As a result, differences between human Achilles tendon and yellow ligament were elucidated at molecular level.

  20. The impact of extracellular matrix coatings on the performance of human renal cells applied in bioartificial kidneys.

    Science.gov (United States)

    Zhang, Huishi; Tasnim, Farah; Ying, Jackie Y; Zink, Daniele

    2009-05-01

    Extracellular matrix (ECM) coatings have been used to improve cell performance in bioartificial kidneys (BAKs). However, their effects on primary human renal proximal tubule cells (HPTCs), which is the most important cell type with regard to clinical applications, have not been tested systematically. Also, the effects of ECM coatings on cell performance during extended time periods have not been addressed. Studying such effects is important for the development of long-term applications. Herein we analyzed for the first time systematically the effects of ECM coatings on proliferation and differentiation of human renal cells and we addressed, in particular, formation and long-term maintenance of differentiated epithelia. Our study focused on HPTCs. ECM coatings were tested alone or in combination with the growth factor bone morphogenetic protein-7 and other additives. The best results were obtained with ECMs consisting of the basal lamina components, laminin or collagen IV, and differentiated epithelia could be maintained up to three weeks on these ECMs. These results provide for the first time clear evidence which kinds of ECM coatings are most appropriate for BAKs. The results also showed that alpha-SMA-expressing myofibroblasts played a key role in the final disruption of differentiated epithelia. This suggests that epithelial-to-mesenchymal transition-related processes might be the major obstacle in long-term applications and such processes should be carefully addressed in future BAK-related research.

  1. Ex-Vivo Tissues Engineering Modeling for Reconstructive Surgery Using Human Adult Adipose Stem Cells and Polymeric Nanostructured Matrix.

    Science.gov (United States)

    Morena, Francesco; Argentati, Chiara; Calzoni, Eleonora; Cordellini, Marino; Emiliani, Carla; D'Angelo, Francesco; Martino, Sabata

    2016-03-31

    The major challenge for stem cell translation regenerative medicine is the regeneration of damaged tissues by creating biological substitutes capable of recapitulating the missing function in the recipient host. Therefore, the current paradigm of tissue engineering strategies is the combination of a selected stem cell type, based on their capability to differentiate toward committed cell lineages, and a biomaterial, that, due to own characteristics (e.g., chemical, electric, mechanical property, nano-topography, and nanostructured molecular components), could serve as active scaffold to generate a bio-hybrid tissue/organ. Thus, effort has been made on the generation of in vitro tissue engineering modeling. Here, we present an in vitro model where human adipose stem cells isolated from lipoaspirate adipose tissue and breast adipose tissue, cultured on polymeric INTEGRA ® Meshed Bilayer Wound Matrix (selected based on conventional clinical applications) are evaluated for their potential application for reconstructive surgery toward bone and adipose tissue. We demonstrated that human adipose stem cells isolated from lipoaspirate and breast tissue have similar stemness properties and are suitable for tissue engineering applications. Finally, the overall results highlighted lipoaspirate adipose tissue as a good source for the generation of adult adipose stem cells.

  2. Comparison of four decontamination treatments on porcine renal decellularized extracellular matrix structure, composition, and support of human renal cortical tubular epithelium cells.

    Science.gov (United States)

    Poornejad, Nafiseh; Nielsen, Jeffery J; Morris, Ryan J; Gassman, Jason R; Reynolds, Paul R; Roeder, Beverly L; Cook, Alonzo D

    2016-03-01

    Engineering whole organs from porcine decellularized extracellular matrix and human cells may lead to a plentiful source of implantable organs. Decontaminating the porcine decellularized extracellular matrix scaffolds is an essential step prior to introducing human cells. However, decontamination of whole porcine kidneys is a major challenge because the decontamination agent or irradiation needs to diffuse deep into the structure to eliminate all microbial contamination while minimizing damage to the structure and composition of the decellularized extracellular matrix. In this study, we compared four decontamination treatments that could be applicable to whole porcine kidneys: 70% ethanol, 0.2% peracetic acid in 1 M NaCl, 0.2% peracetic acid in 4% ethanol, and gamma (γ)-irradiation. Porcine kidneys were decellularized by perfusion of 0.5% (w/v) aqueous solution of sodium dodecyl sulfate and the four decontamination treatments were optimized using segments (n = 60) of renal tissue to ensure a consistent comparison. Although all four methods were successful in decontamination, γ-irradiation was very damaging to collagen fibers and glycosaminoglycans, leading to less proliferation of human renal cortical tubular epithelium cells within the porcine decellularized extracellular matrix. The effectiveness of the other three optimized solution treatments were then all confirmed using whole decellularized porcine kidneys (n = 3). An aqueous solution of 0.2% peracetic acid in 1 M NaCl was determined to be the best method for decontamination of porcine decellularized extracellular matrix. © The Author(s) 2015.

  3. Gallic Acid Inhibited Matrix Invasion and AP-1/ETS-1-Mediated MMP-1 Transcription in Human Nasopharyngeal Carcinoma Cells.

    Science.gov (United States)

    Pang, Jong-Hwei S; Yen, Jia-Hau; Wu, Hsiao-Ting; Huang, Sheng-Teng

    2017-06-24

    Gallic acid is a trihydroxybenzoic acid found in natural herbal plants. Gallic acid has been reported to inhibit the migration and invasive capability of various cancers. Little is known about the underlying mechanisms of invasion responsible for cancer metastasis via gallic acid. The present study was intended to investigate the anti-invasive effect of gallic acid on human nasopharyngeal carcinoma cells (NPC-BM1) and its related mechanism. Gallic acid inhibited the invasion of NPC-BM1 cells dose- and time-dependently without significant cytotoxic effect. Affymetrix oligonucleotide microarray analysis revealed matrix metalloproteinase-1 (MMP-1) as the most down-regulated gene in NPC-BM1 cells by gallic acid. The cytosolic and secreted MMP-1 levels were both found to be inhibited by gallic acid as demonstrated by western blot analysis and ELISA respectively. The mRNA expression and transcription of MMP-1 gene was also down-regulated as determined by RT/real-time PCR and promoter activity assay. The expression of two major transcription binding factors in the MMP-1 promoter, AP-1 and ETS-1, were demonstrated to be reduced by gallic acid in NPC-BM1 cells. The effect of gallic acid was associated with the inhibition of p38 MAPK signaling pathway. In addition, gallic acid enhanced the gene expression of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) which further suppressed the MMP-1 activity. These findings may be useful to develop a novel chemotherapeutic agent to inhibit the metastasis of nasopharyngeal cancer.

  4. Phototherapy up-regulates dentin matrix proteins expression and synthesis by stem cells from human-exfoliated deciduous teeth.

    Science.gov (United States)

    Turrioni, Ana Paula S; Basso, Fernanda G; Montoro, Liege A; Almeida, Leopoldina de Fátima D de; Costa, Carlos A de Souza; Hebling, Josimeri

    2014-10-01

    The aim of this study was to evaluate the effects of infrared LED (850nm) irradiation on dentin matrix proteins expression and synthesis by cultured stem cells from human exfoliated deciduous teeth (SHED). Near-exfoliation primary teeth were extracted (n=3), and SHED cultures were characterized by immunofluorescence using STRO-1, CD44, CD146, Nanog and OCT3/4 antibodies, before experimental protocol. The SHEDs were seeded (3×10(4) cells/cm(2)) with DMEM containing 10% FBS. After 24-h incubation, the culture medium was replaced by osteogenic differentiation medium, and the cells were irradiated with LED light at energy densities (EDs) of 0 (control), 2, or 4J/cm(2) (n=8). The irradiated SHEDs were then evaluated for alkaline phosphatase (ALP) activity, total protein (TP) production, and collagen synthesis (SIRCOL™ Assay), as well as ALP, collagen type I (Col I), dentin sialophosphoprotein (DSPP), and dentin matrix acidic phosphoprotein (DMP-1) gene expression (qPCR). Data were analyzed by Kruskal-Wallis and Mann-Whitney tests (α=0.05). Increased ALP activity and collagen synthesis, as well as gene expression of DSPP and ALP, were observed for both EDs compared with non-irradiated cells. The ED of 4J/cm(2) also increased gene expression of COL I and DMP-1. In conclusion, infrared LED irradiation was capable of biostimulating SHEDs by increasing the expression and synthesis of proteins related with mineralized tissue formation, with overall better results for the energy dose of 4J/cm(2). Phototherapy is an additional approach for the clinical application of LED in Restorative Dentistry. Infrared LED irradiation of the cavity's floor could biostimulate subjacent pulp cells, improving local tissue healing. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Diosgenin, a steroidal saponin, inhibits migration and invasion of human prostate cancer PC-3 cells by reducing matrix metalloproteinases expression.

    Directory of Open Access Journals (Sweden)

    Pin-Shern Chen

    Full Text Available BACKGROUND: Diosgenin, a steroidal saponin obtained from fenugreek (Trigonella foenum graecum, was found to exert anti-carcinogenic properties, such as inhibiting proliferation and inducing apoptosis in a variety of tumor cells. However, the effect of diosgenin on cancer metastasis remains unclear. The aim of the study is to examine the effect of diosgenin on migration and invasion in human prostate cancer PC-3 cells. METHODS AND PRINCIPAL FINDINGS: Diosgenin inhibited proliferation of PC-3 cells in a dose-dependent manner. When treated with non-toxic doses of diosgenin, cell migration and invasion were markedly suppressed by in vitro wound healing assay and Boyden chamber invasion assay, respectively. Furthermore, diosgenin reduced the activities of matrix metalloproteinase-2 (MMP-2 and MMP-9 by gelatin zymography assay. The mRNA level of MMP-2, -9, -7 and extracellular inducer of matrix metalloproteinase (EMMPRIN were also suppressed while tissue inhibitor of metalloproteinase-2 (TIMP-2 was increased by diosgenin. In addition, diosgenin abolished the expression of vascular endothelial growth factor (VEGF in PC-3 cells and tube formation of endothelial cells. Our immunoblotting assays indicated that diosgenin potently suppressed the phosphorylation of phosphatidylinositide-3 kinase (PI3K, Akt, extracellular signal regulating kinase (ERK and c-Jun N-terminal kinase (JNK. In addition, diosgenin significantly decreased the nuclear level of nuclear factor kappa B (NF-κB, suggesting that diosgenin inhibited NF-κB activity. CONCLUSION/SIGNIFICANCE: The results suggested that diosgenin inhibited migration and invasion of PC-3 cells by reducing MMPs expression. It also inhibited ERK, JNK and PI3K/Akt signaling pathways as well as NF-κB activity. These findings reveal new therapeutic potential for diosgenin in anti-metastatic therapy.

  6. Extracellular matrix collagen alters cell proliferation and cell cycle progression of human uterine leiomyoma smooth muscle cells.

    Science.gov (United States)

    Koohestani, Faezeh; Braundmeier, Andrea G; Mahdian, Arash; Seo, Jane; Bi, JiaJia; Nowak, Romana A

    2013-01-01

    Uterine leiomyomas (ULs) are benign tumors occurring in the majority of reproductive aged women. Despite the high prevalence of these tumors, little is known about their etiology. A hallmark of ULs is the excessive deposition of extracellular matrix (ECM), primarily collagens. Collagens are known to modulate cell behavior and function singularly or through interactions with integrins and growth factor-mediated mitogenic pathways. To better understand the pathogenesis of ULs and the role of ECM collagens in their growth, we investigated the interaction of leiomyoma smooth muscle cells (LSMCs) with two different forms of collagen, non-polymerized collagen (monomeric) and polymerized collagen (fibrillar), in the absence or presence of platelet-derived growth factor (PDGF), an abundant growth factor in ULs. Primary cultures of human LSMCS from symptomatic patients were grown on these two different collagen matrices and their morphology, cytoskeletal organization, cellular proliferation, and signaling pathways were evaluated. Our results showed that LSMCs had distinct morphologies on the different collagen matrices and their basal as well as PDGF-stimulated proliferation varied on these matrices. These differences in proliferation were accompanied by changes in cell cycle progression and p21, an inhibitory cell cycle protein. In addition we found alterations in the phosphorylation of focal adhesion kinase, cytoskeletal reorganization, and activation of the mitogen activated protein kinase (MAPK) signaling pathway. In conclusion, our results demonstrate a direct effect of ECM on the proliferation of LSMCs through interplay between the collagen matrix and the PDGF-stimulated MAPK pathway. In addition, these findings will pave the way for identifying novel therapeutic approaches for ULs that target ECM proteins and their signaling pathways in ULs.

  7. CAPN 7 promotes the migration and invasion of human endometrial stromal cell by regulating matrix metalloproteinase 2 activity.

    Science.gov (United States)

    Liu, Hongyu; Jiang, Yue; Jin, Xiaoyan; Zhu, Lihua; Shen, Xiaoyue; Zhang, Qun; Wang, Bin; Wang, Junxia; Hu, Yali; Yan, Guijun; Sun, Haixiang

    2013-07-15

    Matrix metalloproteinase 2 (MMP-2) has been reported to be an important regulator of cell migration and invasion through degradation of the extracellular matrix (ECM) in many diseases, such as cancer and endometriosis. Here, we found calcium-activated neutral protease 7 (CAPN 7) expression was markedly upregulated in the eutopic endometrium and endometrial stromal cells of women diagnosed with endometriosis. Our studies were carried out to detect the effects of CAPN 7 on human endometrial stromal cell (hESC) migration and invasion. Western blotting and quantitative real-time PCR were used to detect the expression of CAPN 7 in endometriosis patients and normal fertile women. Scratch-wound-healing and invasion chamber assay were used to investigate the role of CAPN 7 in hESC migration and invasion. Western blotting, quantitative real-time PCR and zymography were carried out to detect the effect of CAPN 7 on the expressions and activity of MMP-2. CAPN 7 was markedly up-regulated in endometriosis, thereby promoting the migration and invasion of hESC. CAPN 7 overexpression led to increased expression of MMP-2 and tissue inhibitor of metalloproteinases 2 (TIMP-2); CAPN 7 knockdown reversed these changes. CAPN 7 increased MMP-2 activity by increasing the ratio of MMP-2 to TIMP-2. We also found that OA-Hy (an MMP-2 inhibitor) decreased the effects of CAPN 7 overexpression on hESC migration and invasion by approximately 50% and 55%, respectively. Additionally, a coimmunoprecipitation assay demonstrated that CAPN 7 interacted with activator protein 2α (AP-2α): an important transcription factor of MMP-2. CAPN 7 promotes hESC migration and invasion by increasing the activity of MMP-2 via an increased ratio of MMP-2 to TIMP-2.

  8. Effect of apoptosis and response of extracellular matrix proteins after chemotherapy application on human breast cancer cell spheroids.

    Science.gov (United States)

    Oktem, G; Vatansever, S; Ayla, S; Uysal, A; Aktas, S; Karabulut, B; Bilir, A

    2006-02-01

    Multicellular tumor spheroid (MTS) represents a three-dimensional structural form of tumors in laboratory conditions, and it has the characteristics of avascular micrometastases or intervascular spaces of big tumors. Recent studies indicate that extracellular matrix (ECM) proteins play a critical role in tumor metastasis, therefore normal and cancer cells require an ECM for survival, proliferation and differentiation. Doxorubicin and Docetaxel are widely used in the therapy of breast cancer, as well as in in vivo and in vitro studies. In this study, we examined the effect of apoptosis and proliferation of cells on the human breast cancer cell line, MCF-7, by using p53, bcl-2 and Ki67 gene expression, and the tendency to metastasis with extracellular matrix proteins, laminin and type IV collagen after chemotherapy in the spheroid model. The apoptotic cell death in situ was detected by TUNEL method. TUNEL-positive cells and positive immunoreactivities of laminin, type IV collagen, p53 and, bcl-2 were detected in the control group. There was no laminin and type IV collagen immunoreactivities in spheroids of drug groups. While TUNEL-positive cells and p53 immunoreactivity were detected in Docetaxel, Doxorubicin and Docetaxel/Doxorubicin groups, p53 immunoreactivity was not observed in the Docetaxel group. There was no bcl-2 immunoreactivity in either drug group. In addition, we did not detect Ki67 immunoreactivity in both control and drug treatment groups. However, the absence of Ki67 protein in MCF-7 breast multicellular tumor spheroids is possibly related to the cells in G0 or S phase. These chemotherapeutic agents may affect the presence of ECM proteins in this in vitro model of micrometastasis of spheroids. These findings suggest that the possible mechanism of cell death in Doxorubicin and Docetaxel/Doxorubicin treatment groups is related to apoptosis through the p53 pathway. However, we considered the possibility that there is another control mechanism for the

  9. Culturing on decellularized extracellular matrix enhances antioxidant properties of human umbilical cord-derived mesenchymal stem cells

    International Nuclear Information System (INIS)

    Liu, Xiaozhen; Zhou, Long; Chen, Xi; Liu, Tao; Pan, Guoqing; Cui, Wenguo; Li, Mao; Luo, Zong-Ping; Pei, Ming; Yang, Huilin; Gong, Yihong; He, Fan

    2016-01-01

    Human umbilical cord-derived mesenchymal stem cells (UC-MSCs) have attracted great interest in clinical application because of their regenerative potential and their lack of ethical issues. Our previous studies showed that decellularized cell-deposited extracellular matrix (ECM) provided an in vivo-mimicking microenvironment for MSCs and facilitated in vitro cell expansion. This study was conducted to analyze the cellular response of UC-MSCs when culturing on the ECM, including reactive oxygen species (ROS), intracellular antioxidative enzymes, and the resistance to exogenous oxidative stress. After decellularization, the architecture of cell-deposited ECM was characterized as nanofibrous, collagen fibrils and the matrix components were identified as type I and III collagens, fibronectin, and laminin. Compared to tissue culture polystyrene (TCPS) plates, culturing on ECM yielded a 2-fold increase of UC-MSC proliferation and improved the percentage of cells in the S phase by 2.4-fold. The levels of intracellular ROS and hydrogen peroxide (H_2O_2) in ECM-cultured cells were reduced by 41.7% and 82.9%, respectively. More importantly, ECM-cultured UC-MSCs showed enhanced expression and activity of intracellular antioxidative enzymes such as superoxide dismutase and catalase, up-regulated expression of silent information regulator type 1, and suppressed phosphorylation of p38 mitogen-activated protein kinase. Furthermore, a continuous treatment with exogenous 100 μM H_2O_2 dramatically inhibited osteogenic differentiation of UC-MSCs cultured on TCPS, but culturing on ECM retained the differentiation capacity for matrix mineralization and osteoblast-specific marker gene expression. Collectively, by providing sufficient cell amounts and enhancing antioxidant capacity, decellularized ECM can be a promising cell culture platform for in vitro expansion of UC-MSCs. - Highlights: • Decellularization preserved the architecture and components of cell-deposited ECM.

  10. Culturing on decellularized extracellular matrix enhances antioxidant properties of human umbilical cord-derived mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xiaozhen [School of Engineering, Sun Yat-sen University, Guangzhou 510006 (China); Zhou, Long; Chen, Xi [Orthopaedic Institute, Soochow University, Suzhou 215007 (China); Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Suzhou 215006 (China); Liu, Tao [Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Suzhou 215006 (China); Pan, Guoqing; Cui, Wenguo; Li, Mao; Luo, Zong-Ping [Orthopaedic Institute, Soochow University, Suzhou 215007 (China); Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Suzhou 215006 (China); Pei, Ming [Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, WV 26506 (United States); Yang, Huilin [Orthopaedic Institute, Soochow University, Suzhou 215007 (China); Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Suzhou 215006 (China); Gong, Yihong, E-mail: gongyih@mail.sysu.edu.cn [School of Engineering, Sun Yat-sen University, Guangzhou 510006 (China); He, Fan, E-mail: fanhe@suda.edu.cn [Orthopaedic Institute, Soochow University, Suzhou 215007 (China); Department of Orthopaedics, The First Affiliated Hospital of Soochow University, Suzhou 215006 (China)

    2016-04-01

    Human umbilical cord-derived mesenchymal stem cells (UC-MSCs) have attracted great interest in clinical application because of their regenerative potential and their lack of ethical issues. Our previous studies showed that decellularized cell-deposited extracellular matrix (ECM) provided an in vivo-mimicking microenvironment for MSCs and facilitated in vitro cell expansion. This study was conducted to analyze the cellular response of UC-MSCs when culturing on the ECM, including reactive oxygen species (ROS), intracellular antioxidative enzymes, and the resistance to exogenous oxidative stress. After decellularization, the architecture of cell-deposited ECM was characterized as nanofibrous, collagen fibrils and the matrix components were identified as type I and III collagens, fibronectin, and laminin. Compared to tissue culture polystyrene (TCPS) plates, culturing on ECM yielded a 2-fold increase of UC-MSC proliferation and improved the percentage of cells in the S phase by 2.4-fold. The levels of intracellular ROS and hydrogen peroxide (H{sub 2}O{sub 2}) in ECM-cultured cells were reduced by 41.7% and 82.9%, respectively. More importantly, ECM-cultured UC-MSCs showed enhanced expression and activity of intracellular antioxidative enzymes such as superoxide dismutase and catalase, up-regulated expression of silent information regulator type 1, and suppressed phosphorylation of p38 mitogen-activated protein kinase. Furthermore, a continuous treatment with exogenous 100 μM H{sub 2}O{sub 2} dramatically inhibited osteogenic differentiation of UC-MSCs cultured on TCPS, but culturing on ECM retained the differentiation capacity for matrix mineralization and osteoblast-specific marker gene expression. Collectively, by providing sufficient cell amounts and enhancing antioxidant capacity, decellularized ECM can be a promising cell culture platform for in vitro expansion of UC-MSCs. - Highlights: • Decellularization preserved the architecture and components of cell

  11. Three-dimensional culture of human mesenchymal stem cells in a polyethylene terephthalate matrix

    International Nuclear Information System (INIS)

    Cao Yanfen; Li Ding; Shang Chunhua; Wang Jufang; Wang Xiaoning; Yang Shangtian

    2010-01-01

    Polyethylene terephthalate (PET) was used as the scaffold material to support the proliferation of human mesenchymal stem cells (hMSCs). The cells were cultured either statically in multi-wells or in a spinner flask agitated at 80 rpm for up to 20 days. To optimize the cell expansion condition, effects of the initial cell density and basic fibroblast growth factor (bFGF) were examined. During culture, cell growth and metabolism were tested. After 20 days, cells were harvested and surface markers were identified and quantified with flow cytometry. The results showed that hMSCs seeded at the lowest density gave the highest expansion fold. hMSCs grown in porous three-dimensional (3D) matrices displayed significantly different characteristics in terms of their proliferation and metabolism. PET matrices with 3D space could sustain cell proliferation for a long time. In addition, a low concentration (5 ng mL -1 ) of bFGF significantly enhanced the expansion of hMSCs in PET. Cell attachment and distribution in PET matrices were studied with confocal laser microscopy and scanning electron microscopy, which also confirmed cell proliferation. Furthermore, most of the cells in PET matrices were CD29, CD44 and CD105 positive, and CD34, CD45 and CD14 negative, confirming that hMSCs cultured in 3D PET matrices can be expanded and maintained in their undifferentiated state for at least 20 days without subculturing.

  12. Matrix-directed differentiation of human adipose-derived mesenchymal stem cells to dermal-like fibroblasts that produce extracellular matrix.

    Science.gov (United States)

    Sivan, Unnikrishnan; Jayakumar, K; Krishnan, Lissy K

    2016-10-01

    Commercially available skin substitutes lack essential non-immune cells for adequate tissue regeneration of non-healing wounds. A tissue-engineered, patient-specific, dermal substitute could be an attractive option for regenerating chronic wounds, for which adipose-derived mesenchymal stem cells (ADMSCs) could become an autologous source. However, ADMSCs are multipotent in nature and may differentiate into adipocytes, osteocytes and chondrocytes in vitro, and may develop into undesirable tissues upon transplantation. Therefore, ADMSCs committed to the fibroblast lineage could be a better option for in vitro or in vivo skin tissue engineering. The objective of this study was to standardize in vitro culture conditions for ADMSCs differentiation into dermal-like fibroblasts which can synthesize extracellular matrix (ECM) proteins. Biomimetic matrix composite, deposited on tissue culture polystyrene (TCPS), and differentiation medium (DM), supplemented with fibroblast-conditioned medium and growth factors, were used as a fibroblast-specific niche (FSN) for cell culture. For controls, ADMSCs were cultured on bare TCPS with either DM or basal medium (BM). Culture of ADMSCs on FSN upregulated the expression of differentiation markers such as fibroblast-specific protein-1 (FSP-1) and a panel of ECM molecules specific to the dermis, such as fibrillin-1, collagen I, collagen IV and elastin. Immunostaining showed the deposition of dermal-specific ECM, which was significantly higher in FSN compared to control. Fibroblasts derived from ADMSCs can synthesize elastin, which is an added advantage for successful skin tissue engineering as compared to fibroblasts from skin biopsy. To obtain rapid differentiation of ADMSCs to dermal-like fibroblasts for regenerative medicine, a matrix-directed differentiation strategy may be employed. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

  13. Extracellular Matrix Hydrogel Derived from Human Umbilical Cord as a Scaffold for Neural Tissue Repair and Its Comparison with Extracellular Matrix from Porcine Tissues

    Czech Academy of Sciences Publication Activity Database

    Kočí, Zuzana; Výborný, Karel; Dubišová, Jana; Vacková, Irena; Jäger, Aleš; Lunov, Oleg; Jiráková, Klára; Kubinová, Šárka

    2017-01-01

    Roč. 23, č. 6 (2017), s. 333-345 ISSN 1937-3384 R&D Projects: GA ČR(CZ) GA15-01396S; GA MŠk(CZ) LO1309; GA MŠk(CZ) ED1.1.00/02.0109; GA MŠk(CZ) EF15_003/0000419 Grant - others:AV ČR(CZ) Fellowship J. E. Purkyně Institutional support: RVO:68378041 ; RVO:68378271 Keywords : extracellular matrix * hydrogel * umbilical cord Subject RIV: FH - Neurology; EB - Genetics ; Molecular Biology (FZU-D) OBOR OECD: Neurosciences (including psychophysiology; Biophysics (FZU-D)

  14. Fibrin network pattern changes of platelet-rich fibrin in young versus old age group of individuals: A cell block cytology study

    OpenAIRE

    Shravanthi Raghav Yajamanya; Anirban Chatterjee; Chaitanya Nischay Babu; Deepika Karunanithi

    2016-01-01

    Background: To evaluate variations in fibrin network patterns of the platelet-rich fibrin (PRF) in different age groups. Materials and Methods: Ninety-five patients were divided into three age groups: Group 1: (20?39 years); Group 2: (40?59 years); and Group 3: (60 years and above). PRF was prepared from blood samples of all patients and were subjected to cell block cytology method of histological analysis and slides were prepared to histologically assess the age-related changes in (i) fibrin...

  15. Fibrin-Enhanced Canonical Wnt Signaling Directs Plasminogen Expression in Cementoblasts

    Directory of Open Access Journals (Sweden)

    Saeed Ur Rahman

    2017-11-01

    Full Text Available Cementum is a mineralized layer on the tooth’s root surface and facilitates the biomechanical anchoring of fibrous connective tissues as a part of tooth-supportive complexes. Previously, we observed that OCCM30 cementoblasts cultured on fibrin matrices underwent apoptosis due to fibrin degradation through the expression of proteases. Here, we demonstrated that OCCM30 on fibrin matrices (OCCM30-fibrin enhanced canonical Wnt signaling, which directed to plasminogen expression. The OCCM30-fibrin showed higher levels of Wnt3a expression, nuclear translocation of β-catenin, and T-cell factor (TCF optimal motif (TOP reporter activity than the cells on tissue culture dishes (OCCM30-TCD, indicating that the OCCM30-fibrin enhanced canonical Wnt/β-catenin signaling. Also, OCCM30-fibrin expressed biomineralization-associated markers at higher levels than OCCM30-TCD, of which levels were further increased with LiCl, a Wnt signaling activator. The OCCM30 cementoblasts simultaneously showed that high levels of plasminogen, a critical component of fibrinolysis, were expressed in the OCCM30-fibrin. Activation of canonical Wnt signaling with LiCl treatment or with forced lymphoid enhancer factor 1 (LEF1-expression increased the expression of plasminogen. On the contrary, the inhibition of canonical Wnt signaling with siRNAs against Wnt3a or β-catenin abrogated fibrin-enhanced plasminogen expression. Furthermore, there are three conserved putative response elements for the LEF1/β-catenin complex in the plasminogen proximal promoter regions (−900 to +54. Site-directed mutations and chromatin immunoprecipitation indicated that canonical Wnt signaling directed plasminogen expression. Taken together, this study suggests that fibrin-based materials can modulate functional periodontal formations in controlling cementoblast differentiation and fibrin degradation.

  16. Ex Vivo and In Vivo Imaging and Biodistribution of Aptamers Targeting the Human Matrix MetalloProtease-9 in Melanomas.

    Directory of Open Access Journals (Sweden)

    David Kryza

    Full Text Available The human Matrix MetalloProtease-9 (hMMP-9 is overexpressed in tumors where it promotes the release of cancer cells thus contributing to tumor metastasis. We raised aptamers against hMMP-9, which constitutes a validated marker of malignant tumors, in order to design probes for imaging tumors in human beings. A chemically modified RNA aptamer (F3B, fully resistant to nucleases was previously described. This compound was subsequently used for the preparation of F3B-Cy5, F3B-S-acetylmercaptoacetyltriglycine (MAG and F3B-DOTA. The binding properties of these derivatives were determined by surface plasmon resonance and electrophoretic mobility shift assay. Optical fluorescence imaging confirmed the binding to hMMP-9 in A375 melanoma bearing mice. Quantitative biodistribution studies were performed at 30 min, 1h and 2 h post injection of 99mTc-MAG-aptamer and 111In-DOTA-F3B. 99mTc radiolabeled aptamer specifically detected hMMP-9 in A375 melanoma tumors but accumulation in digestive tract was very high. Following i.v. injection of 111In-DOTA-F3B, high level of radioactivity was observed in kidneys and bladder but digestive tract uptake was very limited. Tumor uptake was significantly (student t test, p<0.05 higher for 111In-DOTA-F3B with 2.0%ID/g than for the 111In-DOTA-control oligonucleotide (0.7%ID/g with tumor to muscle ratio of 4.0. Such difference in tumor accumulation has been confirmed by ex vivo scintigraphic images performed at 1h post injection and by autoradiography, which revealed the overexpression of hMMP-9 in sections of human melanomas. These results demonstrate that F3B aptamer is of interest for detecting hMMP-9 in melanoma tumor.

  17. Plasminogen activator activity and plasma-coagulum lysis measured by use of optimized fibrin gel structure preformed in microtiter plates

    DEFF Research Database (Denmark)

    Sidelmann, Johannes Jakobsen; Jespersen, J; Gram, J

    1995-01-01

    We introduce a new fibrin plate assay performed in microtiter plates. By means of spectroscopic studies we optimized the structure of the fibrin gel and then used the optimized fibrin gel to determine plasminogen activator activity. Plasminogen activator solutions were applied on top of the fibri...

  18. Bioactivity and stability of endogenous fibrogenic factors in platelet-rich fibrin

    DEFF Research Database (Denmark)

    Lundquist, R.; Dziegiel, M.H.; Agren, M.S.

    2008-01-01

    Platelet-rich fibrin (PRF) is an autologous fibrin sealant (FS) enriched with a platelet concentrate (> 1,000,000 platelets/microL) produced by the automated Vivostat system and used to enhance wound healing. The effects of PRF were compared with supernatant from thrombin-activated platelet conce...

  19. Anti-fibrin antibody binding in valvular vegetations and kidney lesions during experimental endocarditis.

    Science.gov (United States)

    Yokota, M; Basi, D L; Herzberg, M C; Meyer, M W

    2001-01-01

    In Streptococcus sanguinis (sanguis) induced experimental endocarditis, we sought evidence that the development of aortic valvular vegetation depends on the availability of fibrin. Endocarditis was induced in New Zealand white rabbits by catheter placement into the left ventricle and inoculation of the bacteria. Fibrin was localized in the developing vegetation with 99mTechnetium (Tc)-labeled anti-fibrin antibody one or three days later. When rabbit anti-fibrin antibody was given intravenously on day 1, the mass of aortic valvular vegetation was significantly reduced at day 3; infusion of non-specific rabbit IgG showed no effect. The 99mTc-labeled anti-fibrin antibody also labeled kidneys that showed macroscopic subcapsular hemorrhage. To learn if the deposition of fibrin in the kidneys was a consequence of endocarditis required a comparison of farm-bred and specific pathogen-free rabbits before and after the induction of endocarditis. Before induction, the kidneys of farm-bred rabbits were labeled, but specific pathogen-free rabbits were free of labeling and signs of macroscopic hemorrhage. After 3 days of endocarditis, kidneys of 10 of 14 specific pathogen-free rabbits labeled with 99mTc-labeled anti-fibrin antibody and showed hemorrhage. Kidney lesions were suggested to be a frequent sequellae of S. sanguinis infective endocarditis. For the first time, fibrin was shown to be required for the continued development of aortic valvular vegetations.

  20. Disruption of fibronectin matrix affects type IV collagen, fibrillin and laminin deposition into extracellular matrix of human trabecular meshwork (HTM) cells.

    Science.gov (United States)

    Filla, Mark S; Dimeo, Kaylee D; Tong, Tiegang; Peters, Donna M

    2017-12-01

    Fibronectin fibrils are a major component of the extracellular matrix (ECM) of the trabecular meshwork (TM). They are a key mediator of the formation of the ECM which controls aqueous humor outflow and contributes to the pathogenesis of glaucoma. The purpose of this work was to determine if a fibronectin-binding peptide called FUD, derived from the Streptococcus pyogenes Functional Upstream Domain of the F1 adhesin protein, could be used to control fibronectin fibrillogenesis and hence ECM formation under conditions where its expression was induced by treatment with the glucocorticoid dexamethasone. FUD was very effective at preventing fibronectin fibrillogenesis in the presence or absence of steroid treatment as well as the removal of existing fibronectin fibrils. Disruption of fibronectin fibrillogenesis by FUD also disrupted the incorporation of type IV collagen, laminin and fibrillin into the ECM. The effect of FUD on these other protein matrices, however, was found to be dependent upon the maturity of the ECM when FUD was added. FUD effectively disrupted the incorporation of these other proteins into matrices when added to newly confluent cells that were forming a nascent ECM. In contrast, FUD had no effect on these other protein matrices if the cell cultures already possessed a pre-formed, mature ECM. Our studies indicate that FUD can be used to control fibronectin fibrillogenesis and that these fibrils play a role in regulating the assembly of other ECM protein into matrices involving type IV collagen, laminin, and fibrillin within the TM. This suggests that under in vivo conditions, FUD would selectively disrupt fibronectin fibrils and de novo assembly of other proteins into the ECM. Finally, our studies suggest that targeting fibronectin fibril assembly may be a viable treatment for POAG as well as other glaucomas involving excessive or abnormal matrix deposition of the ECM. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Structure and function of ameloblastin as an extracellular matrix protein: adhesion, calcium binding, and CD63 interaction in human and mouse.

    Science.gov (United States)

    Zhang, Xu; Diekwisch, Thomas G H; Luan, Xianghong

    2011-12-01

    The functional significance of extracellular matrix proteins in the life of vertebrates is underscored by a high level of sequence variability in tandem with a substantial degree of conservation in terms of cell-cell and cell-matrix adhesion interactions. Many extracellular matrix proteins feature multiple adhesion domains for successful attachment to substrates, such as integrin, CD63, and heparin. Here we have used homology and ab initio modeling algorithms to compare mouse ameloblastin (mAMBN) and human ameloblastin (hABMN) isoforms and to analyze their potential for cell adhesion and interaction with other matrix molecules as well as calcium binding. Sequence comparison between mAMBN and hAMBN revealed a 26-amino-acid deletion in mAMBN, corresponding to a helix-loop-helix frameshift. The human AMBN domain (174Q-201G), homologous to the mAMBN 157E-178I helix-loop-helix region, formed a helix-loop motif with an extended loop, suggesting a higher degree of flexibility of hAMBN compared with mAMBN, as confirmed by molecular dynamics simulation. Heparin-binding domains, CD63-interaction domains, and calcium-binding sites in both hAMBN and mAMBN support the concept of AMBN as an extracellular matrix protein. The high level of conservation between AMBN functional domains related to adhesion and differentiation was remarkable when compared with only 61% amino acid sequence homology. © 2011 Eur J Oral Sci.

  2. Eicosapentaenoic acid inhibits TNF-α-induced matrix metalloproteinase-9 expression in human keratinocytes, HaCaT cells

    International Nuclear Information System (INIS)

    Kim, Hyeon Ho; Lee, Youngae; Eun, Hee Chul; Chung, Jin Ho

    2008-01-01

    Eicosapentaenoic acid (EPA) is an omega-3 (ω-3) polyunsaturated fatty acid (PUFA), which has anti-inflammatory and anti-cancer properties. Some reports have demonstrated that EPA inhibits NF-κB activation induced by tumor necrosis factor (TNF)-α or lipopolysaccharide (LPS) in various cells. However, its detailed mode of action is unclear. In this report, we investigated whether EPA inhibits the expression of TNF-α-induced matrix metalloproteinases (MMP)-9 in human immortalized keratinocytes (HaCaT). TNF-α induced MMP-9 expression by NF-κB-dependent pathway. Pretreatment of EPA inhibited TNF-α-induced MMP-9 expression and p65 phosphorylation. However, EPA could not affect IκB-α phosphorylation, nuclear translocation of p65, and DNA binding activity of NF-κB. EPA inhibited TNF-α-induced p65 phosphorylation through p38 and Akt inhibition and this inhibition was IKKα-dependent event. Taken together, we demonstrate that EPA inhibits TNF-α-induced MMP-9 expression through inhibition of p38 and Akt activation

  3. Rapid Quantification of 25-Hydroxyvitamin D3 in Human Serum by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

    Science.gov (United States)

    Qi, Yulin; Müller, Miriam; Stokes, Caroline S.; Volmer, Dietrich A.

    2018-04-01

    LC-MS/MS is widely utilized today for quantification of vitamin D in biological fluids. Mass spectrometric assays for vitamin D require very careful method optimization for precise and interference-free, accurate analyses however. Here, we explore chemical derivatization and matrix-assisted laser desorption/ionization (MALDI) as a rapid alternative for quantitative measurement of 25-hydroxyvitamin D3 in human serum, and compare it to results from LC-MS/MS. The method implemented an automated imaging step of each MALDI spot, to locate areas of high intensity, avoid sweet spot phenomena, and thus improve precision. There was no statistically significant difference in vitamin D quantification between the MALDI-MS/MS and LC-MS/MS: mean ± standard deviation for MALDI-MS—29.4 ± 10.3 ng/mL—versus LC-MS/MS—30.3 ± 11.2 ng/mL (P = 0.128)—for the sum of the 25-hydroxyvitamin D epimers. The MALDI-based assay avoided time-consuming chromatographic separation steps and was thus much faster than the LC-MS/MS assay. It also consumed less sample, required no organic solvents, and was readily automated. In this proof-of-concept study, MALDI-MS readily demonstrated its potential for mass spectrometric quantification of vitamin D compounds in biological fluids.

  4. Outcome of nonunion fractures in dogs treated with fixation, compression resistant matrix, and recombinant human bone morphogenetic protein-2.

    Science.gov (United States)

    Massie, Anna M; Kapatkin, Amy S; Fuller, Mark C; Verstraete, Frank J M; Arzi, Boaz

    2017-03-20

    To report the use of compression resistant matrix (CRM) infused with recombinant human bone morphogenetic protein (rhBMP-2) prospectively in the healing of nonunion long-bone fractures in dogs. A longitudinal cohort of dogs that were presented with nonunion fractures were classified and treated with CRM soaked with rhBMP-2 and fracture fixation. They were followed with serial radiographs and evaluated for healing times and complications according to the time frame and definitions previously established for orthopaedic clinical cases. Eleven nonunion fractures in nine dogs were included. Median healing time was 10 weeks (range: 7-20 weeks). Major perioperative complications due to bandage morbidity were encountered in two of 11 limbs and resolved. All other complications were minor. They occurred perioperatively in eight of 11 limbs. Minor follow-up complications included short-term in one of two limbs, mid-term in one of three, and long-term in four of five limbs. Nine limbs returned to full function and two limbs returned to acceptable function at the last follow-up. Nonunion fractures given a poor prognosis via standard-of-care treatment were successfully repaired using CRM with rhBMP-2 accompanying fixation. These dogs, previously at high risk of failure, returned to full or acceptable function.

  5. New low-flux mixed matrix membranes that offer superior removal of protein-bound toxins from human plasma

    Science.gov (United States)

    Pavlenko, Denys; van Geffen, Esmée; van Steenbergen, Mies J.; Glorieux, Griet; Vanholder, Raymond; Gerritsen, Karin G. F.; Stamatialis, Dimitrios

    2016-10-01

    Hemodialysis is a widely available and well-established treatment for patients with End Stage Renal Disease (ESRD). However, although life-sustaining, patient mortality rates are very high. Several recent studies corroborated the link between dialysis patients’ outcomes and elevated levels of protein-bound uremic toxins (PBUT) that are poorly removed by conventional hemodialysis. Therefore, new treatments are needed to improve their removal. Recently, our group showed that the combination of dialysis and adsorption on one membrane, the mixed matrix membrane (MMM), can effectively remove those toxins from human plasma. However, these first MMMs were rather large in diameter and their mass transport characteristics needed improvement before application in the clinical setting. Therefore, in this study we developed a new generation of MMMs that have a smaller diameter and optimized characteristics offering superior ability in removing the PBUT indoxyl sulfate (IS) and p-cresyl sulfate (pCS) in comparison to first generation MMMs (30 and 125% respectively), as well as, a commercial dialysis membrane (more than 100% better removal).

  6. Demineralised human dentine matrix stimulates the expression of VEGF and accelerates the bone repair in tooth sockets of rats.

    Science.gov (United States)

    Reis-Filho, Cláudio R; Silva, Elisângela R; Martins, Adalberto B; Pessoa, Fernanda F; Gomes, Paula V N; de Araújo, Mariana S C; Miziara, Melissa N; Alves, José B

    2012-05-01

    In this study we investigated the possible use of human demineralised dentine matrix (DHDM), obtained from the extracted teeth, as bone graft material and evaluated the expression of vascular endothelial growth factor (VEGF) induced by this material in the healing process of tooth sockets of rats. To evaluate bone regeneration and expression of VEGF induced by DHDM, thirty-two male Wistar rats weighing approximately 200 g were used. After maxillary second molar extraction, the left sockets were filled with DHDM and the right sockets were naturally filled by blood clot (control). The animals were sacrificed at 3, 7, 14 and 21 days after surgery and upper maxillaries were processed for histological, morphometric and immunohistochemical analyses. DHDM was used to evaluate the mechanical effect of bone graft material into sockets. Expression of VEGF was determined by immunohistochemistry in all groups. Our results demonstrated a significant increase in the newly formed bone tissue in sockets of 7, 14 and 21 days and a significant increase in VEGF expression at days 7 and 14 on treated sockets. Our results showed that DHDM increases the expression of VEGF and accelerates the healing process in rats tooth sockets, by stimulating bone deposition and also vessels formation. These results suggest that DHDM has osteoinductive/osteoconductive potential and may represent an efficient grafting material on guided bone regeneration. Copyright © 2011 Elsevier Ltd. All rights reserved.

  7. Influence of substrate composition on human embryonic stem cell differentiation and extracellular matrix production in embryoid bodies.

    Science.gov (United States)

    Laperle, Alex; Masters, Kristyn S; Palecek, Sean P

    2015-01-01

    Stem cells reside in specialized niches in vivo. Specific factors, including the extracellular matrix (ECM), in these niches are directly responsible for maintaining the stem cell population. During development, components of the stem cell microenvironment also control differentiation with precise spatial and temporal organization. The stem cell microenvironment is dynamically regulated by the cellular component, including stem cells themselves. Thus, a mechanism exists whereby stem cells modify the ECM, which in turn affects the fate of the stem cell. In this study, we investigated whether the type of ECM initially adsorbed to the culture substrate can influence the composition of the ECM deposited by human embryonic stem cells (hESCs) differentiating in embryoid bodies, and whether different ECM composition and deposition profiles elicit distinct differentiation fates. We have shown that the initial ECM environment hESCs are exposed to affects the fate decisions of those cells and that this initial ECM environment is constantly modified during the differentiation process. © 2014 American Institute of Chemical Engineers.

  8. Matrix metalloproteinase-1 inhibitory activities of Morinda citrifolia seed extract and its constituents in UVA-irradiated human dermal fibroblasts.

    Science.gov (United States)

    Masuda, Megumi; Murata, Kazuya; Naruto, Shunsuke; Uwaya, Akemi; Isami, Fumiyuki; Matsuda, Hideaki

    2012-01-01

    The objective of this study was to examine whether a 50% ethanolic extract (MCS-ext) of the seeds of Morinda citrifolia (noni) and its constituents have matrix metalloproteinase-1 (MMP-1) inhibitory activity in UVA-irradiated normal human dermal fibroblasts (NHDFs). The MCS-ext (10 μg/mL) inhibited MMP-1 secretion from UVA-irradiated NHDFs, without cytotoxic effects, at 48 h after UV exposure. The ethyl acetate-soluble fraction of MCS-ext was the most potent inhibitor of MMP-1 secretion. Among the constituents of the fraction, a lignan, 3,3'-bisdemethylpinoresinol (1), inhibited the MMP-1 secretion at a concentration of 0.3 μM without cytotoxic effects. Furthermore, 1 (0.3 μM) reduced the level of intracellular MMP-1 expression. Other constituents, namely americanin A (2), quercetin (3) and ursolic acid (4), were inactive. To elucidate inhibition mechanisms of MMP-1 expression and secretion, the effect of 1 on mitogen-activated protein kinases (MAPKs) phosphorylation was examined. Western blot analysis revealed that 1 (0.3 μM) reduced the phosphorylations of p38 and c-Jun-N-terminal kinase (JNK). These results suggested that 1 suppresses intracellular MMP-1 expression, and consequent secretion from UVA-irradiated NHDFs, by down-regulation of MAPKs phosphorylation.

  9. Integrin-mediated adhesion of human mesenchymal stem cells to extracellular matrix proteins adsorbed to polymer surfaces

    International Nuclear Information System (INIS)

    Dånmark, S; Mustafa, K; Finne-Wistrand, A; Albertsson, A-C; Patarroyo, M

    2012-01-01

    In vitro, degradable aliphatic polyesters are widely used as cell carriers for bone tissue engineering, despite their lack of biological cues. Their biological active surface is rather determined by an adsorbed layer of proteins from the surrounding media. Initial cell fate, including adhesion and proliferation, which are key properties for efficient cell carriers, is determined by the adsorbed layer of proteins. Herein we have investigated the ability of human bone marrow derived stem cells (hBMSC) to adhere to extracellular matrix (ECM) proteins, including fibronectin and vitronectin which are present in plasma and serum. hBMSC expressed integrins for collagens, laminins, fibronectin and vitronectin. Accordingly, hBMSC strongly adhered to these purified ECM proteins by using the corresponding integrins. Although purified fibronectin and vitronectin adsorbed to aliphatic polyesters to a lower extent than to cell culture polystyrene, these low levels were sufficient to mediate adhesion of hBMSC. It was found that plasma- and serum-coated polystyrene adsorbed significant levels of both fibronectin and vitronectin, and fibronectin was identified as the major adhesive component of plasma for hBMSC; however, aliphatic polyesters adsorbed minimal levels of fibronectin under similar conditions resulting in impaired cell adhesion. Altogether, the results suggest that the efficiency of aliphatic polyesters cell carriers could be improved by increasing their ability to adsorb fibronectin. (paper)

  10. Construction and characterization of human oral mucosa equivalent using hyper-dry amniotic membrane as a matrix.

    Science.gov (United States)

    Qi, Fangfang; Yoshida, Toshiko; Koike, Takeshi; Aizawa, Hitoshi; Shimane, Tetsu; Li, Yinghui; Yamada, Shinichi; Okabe, Motonori; Nikaido, Toshio; Kurita, Hiroshi

    2016-05-01

    Human amniotic membrane(HAM) as a graft material has been used in various fields. Hyper-dry amniotic membrane (HD-AM) is a novel dried amniotic membrane that is easy to handle and can be preserved at room temperature without time limitation. The purpose of this study was to investigate the useful properties of HD-AM in reconstruction of the oral mucosa. Human oral keratinocytes were isolated and seeded on HD-AM in serum-free culture system. Oral mucosa equivalent (OME) was developed and transplanted onto full-thickness wound on athymic mice. The wound healing was analyzed and the OME both before and after transplantation was analyzed with hematoxylin-eosin staining and immunohistochemical staining for Cytokines 10 (CK10), Cytokines 16 (CK16), and Ivolucrin (IVL). Oral keratinocytes spread and proliferated well on HD-AM. Two weeks after air-lifting, OME had formed with good differentiation and morphology. We confirmed immunohistochemically that the expression of CK10 was positive in all suprabasal layers, as was CK16 in the upper layers, while IVL was present in all cell layers. Three weeks after transplantation to athymic mice, the newly generated tissue had survived well with the smallest contraction. The epithelial cells of newly generated tissue expressed CK10 throughout in all suprabasal layers, IVL was mainly in the granular layer, and CK16 positive cells were observed in all spinous layer and granular layer but were not expressed in the mouse skin, all of which were similar to native gingival mucosa. The OME with HD-AM as a matrix revealed a good morphology and stable wound healing. This study demonstrates that HD-AM is a useful and feasible biomaterial for oral mucosa reconstruction. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Quantitative serology assays for determination of antibody responses to Ebola virus glycoprotein and matrix protein in nonhuman primates and humans.

    Science.gov (United States)

    Vu, Hong; Shulenin, Sergey; Grolla, Allen; Audet, Jonathan; He, Shihua; Kobinger, Gary; Unfer, Robert C; Warfield, Kelly L; Aman, M Javad; Holtsberg, Frederick W

    2016-02-01

    The West Africa Ebola virus disease (EVD) outbreak has reached unprecedented magnitude and caused worldwide concerns for the spread of this deadly virus. Recent findings in nonhuman primates (NHPs) demonstrate that antibodies can be protective against EVD. However, the role of antibody response in vaccine-mediated protection is not fully understood. To address these questions quantitative serology assays are needed for measurement of the antibody response to key Ebola virus (EBOV) proteins. Serology enzyme-linked immunosorbent assays (ELISA's), using a reference detection antibody, were developed in order to standardize the quantitation of antibody levels in vaccinated NHPs or in humans exposed to EBOV or immunized with an EBOV vaccine. Critical reagents were generated to support the development of the serology ELISAs. Recombinant EBOV matrix protein (VP40) was expressed in Escherichia coli and purified. Two variants of the glycoprotein (GP), the ectodomain lacking the transmembrane domain (GPΔTM), and an engineered GP lacking the mucin-like domain (GPΔmuc) were expressed and purified from mammalian cell systems. Using these proteins, three ELISA methods were developed and optimized for reproducibility and robustness, including stability testing of critical reagents. The assay was used to determine the antibody response against VP40, GPΔTM, and GPΔmuc in a NHP vaccine study using EBOV virus-like particles (VLP) vaccine expressing GP, VP40 and the nucleoprotein. Additionally, these ELISAs were used to successfully detect antibody responses to VP40, GPΔTM and GPΔmuc in human sera from EBOV infected individuals. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. 10-Hydroxy-2-decenoic acid prevents ultraviolet A-induced damage and matrix metalloproteinases expression in human dermal fibroblasts.

    Science.gov (United States)

    Zheng, Jinfen; Lai, Wei; Zhu, Guoxing; Wan, Miaojian; Chen, Jian; Tai, Yan; Lu, Chun

    2013-10-01

    10-Hydroxy-2-decenoic acid (10-HDA) is a major fatty acid component of royal jelly, which has been reported to have a variety of beneficial pharmacological characteristics. However, the effects of 10-HDA on skin photoageing and its potential mechanism of action are unclear. We investigated the protective effects of 10-HDA on ultraviolet (UV) A-induced damage in human dermal fibroblasts (HDFs). We then explored the inhibitory effects of 10-HDA on UVA-induced matrix metalloproteinases (MMPs) expression and elucidated the signalling pathways controlling MMPs inhibition. Primary human dermal fibroblasts were exposed to UVA. Cell proliferation, cellular senescent state and collagen content were analysed using CCK-8, senescence-associated β-galactosidase staining and Sircol collagen assay, respectively. Fluorometric assays were performed to detect the formation of reactive oxygen species (ROS) in the cells. The mRNA levels of MMP-1, MMP-3 and type I (α1) collagen were determined by quantitative real-time PCR. Western blot was applied to detect the expression of MMP-1, MMP-3, JNK and p38 MAPK. HDFs treated with 10-HDA were significantly protected from UVA-induced cytotoxicity, ROS, cellular senescence and stimulated collagen production. Moreover, 10-HDA suppressed the UVA-induced expression of MMP-1 and MMP-3 at both the transcriptional and protein levels. Treatment with 10-HDA also reduced the UVA-induced activation of the JNK and p38 MAPK pathways. The data obtained in this study provide evidence that 10-HDA could prevent UVA-induced damage and inhibit MMP-1 and MMP-3 expressions. Therefore, 10-HDA may be a potential agent for the prevention and treatment of skin photoageing. © 2012 The Authors. Journal of the European Academy of Dermatology and Venereology © 2012 European Academy of Dermatology and Venereology.

  13. Stiffness of hyaluronic acid gels containing liver extracellular matrix supports human hepatocyte function and alters cell morphology.

    Science.gov (United States)

    Deegan, Daniel B; Zimmerman, Cynthia; Skardal, Aleksander; Atala, Anthony; Shupe, Thomas D

    2015-03-01

    Tissue engineering and cell based liver therapies have utilized primary hepatocytes with limited success due to the failure of hepatocytes to maintain their phenotype in vitro. In order to overcome this challenge, hyaluronic acid (HA) cell culture substrates were formulated to closely mimic the composition and stiffness of the normal liver cellular microenvironment. The stiffness of the substrate was modulated by adjusting HA hydrogel crosslinking. Additionally, the repertoire of bioactive molecules within the HA substrate was bolstered by supplementation with normal liver extracellular matrix (ECM). Primary human hepatocyte viability and phenotype were determined over a narrow physiologically relevant range of substrate stiffnesses from 600 to 4600Pa in both the presence and absence of liver ECM. Cell attachment, viability, and organization of the actin cytoskeleton improved with increased stiffness up to 4600Pa. These differences were not evident in earlier time points or substrates containing only HA. However, gene expression for the hepatocyte markers hepatocyte nuclear factor 4 alpha (HNF4α) and albumin significantly decreased on the 4600Pa stiffness at day 7 indicating that cells may not have maintained their phenotype long-term at this stiffness. Function, as measured by albumin secretion, varied with both stiffness and time in culture and peaked at day 7 at the 1200Pa stiffness, slightly below the stiffness of normal liver ECM at 3000Pa. Overall, gel stiffness affected primary human hepatocyte cell adhesion, functional marker expression, and morphological characteristics dependent on both the presence of liver ECM in gel substrates and time in culture. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Topical photodynamic therapy following excisional wounding of human skin increases production of transforming growth factor-β3 and matrix metalloproteinases 1 and 9, with associated improvement in dermal matrix organization.

    Science.gov (United States)

    Mills, S J; Farrar, M D; Ashcroft, G S; Griffiths, C E M; Hardman, M J; Rhodes, L E

    2014-07-01

    Animal studies report photodynamic therapy (PDT) to improve healing of excisional wounds; the mechanism is uncertain and equivalent human studies are lacking. To explore the impact of methyl aminolaevulinate (MAL)-PDT on clinical and microscopic parameters of human cutaneous excisional wound healing, examining potential modulation through production of transforming growth factor (TGF)-β isoforms. In 27 healthy older men (60-77 years), a 4-mm punch biopsy wound was created in skin of the upper inner arm and treated with MAL-PDT three times over 5 days. An identical control wound to the contralateral arm was untreated and both wounds left to heal by secondary intention. Wounds were re-excised during the inflammatory phase (7 days, n = 10), matrix remodelling (3 weeks, n = 8) and cosmetic outcome/dermal structure (9 months, n = 9). Production of TGF-β1, TGF-β3 and matrix metalloproteinases (MMPs) was assessed by immunohistochemistry alongside microscopic measurement of wound size/area and clinical assessment of wound appearance. MAL-PDT delayed re-epithelialization at 7 days, associated with increased inflammation. However, 3 weeks postwounding, treated wounds were smaller with higher production of MMP-1 (P = 0·01), MMP-9 (P = 0·04) and TGF-β3 (P = 0·03). TGF-β1 was lower than control at 7 days and higher at 3 weeks (both P = 0·03). At 9 months, MAL-PDT-treated wounds showed greater, more ordered deposition of collagen I, collagen III and elastin (all P < 0·05). MAL-PDT increases MMP-1, MMP-9 and TGF-β3 production during matrix remodelling, ultimately producing scars with improved dermal matrix architecture. © 2014 British Association of Dermatologists.

  15. The role of titanium-prepared platelet-rich fibrin in palatal mucosal wound healing and histoconduction.

    Science.gov (United States)

    Ustaoğlu, Gülbahar; Ercan, Esra; Tunali, Mustafa

    2016-10-01

    We aimed to determine the clinical effects of titanium-prepared, platelet-rich fibrin (T-PRF) on human palatal mucosal wound healing (PMWH) and to identify its effect on time-dependent changes in palatal soft-tissue thickness (PSTT) in terms of histoconduction, which is a new concept. Free gingival graft (FGG) donor sites were treated with T-PRF and compared with an untreated control group. The results of colour match and H 2 O 2 -bubbling tests for complete wound epithelization (CWE) were recorded on days 3, 7, 14 and 21. Pain level, number of analgesics used and bleeding status were recorded for the first 7 days. PSTT was measured at baseline and after 1 and 6 months. Colour match scores of the test group were significantly higher than those of the control group at 7 and 14 days. CWE was observed at a higher frequency in the test group than in the control group on day 14. Post-operative bleeding prevalence was lower in the test group than in the control group for the first 2 days. A time-dependent decrease in PSTT was observed at 1 and 6 months in the control group compared with baseline (baseline, 4.23 ± 0.62 mm; 1 month, 4.01 ± 0.68 mm; and 6 months, 3.93 ± 0.69 mm). However, no significant difference was found in the test group (baseline, 4.29 ± 0.64 mm; 1 month, 4.61 ± 0.51 mm; and 6 months, 4.51 ± 0.58 mm). The T-PRF membrane exhibited positive effects on PMWH. T-PRF, which is a promising autogenous matrix for histoconduction, may also be preferred as an autogenous alternative to connective tissue grafts in the treatment of gingival recessions and peri-implant mucosal recessions.

  16. Ultrastructure and growth factor content of equine platelet-rich fibrin gels.

    Science.gov (United States)

    Textor, Jamie A; Murphy, Kaitlin C; Leach, J Kent; Tablin, Fern

    2014-04-01

    To compare fiber diameter, pore area, compressive stiffness, gelation properties, and selected growth factor content of platelet-rich fibrin gels (PRFGs) and conventional fibrin gels (FGs). PRFGs and conventional FGs prepared from the blood of 10 healthy horses. Autologous fibrinogen was used to form conventional FGs. The PRFGs were formed from autologous platelet-rich plasma of various platelet concentrations (100 × 10³ platelets/μL, 250 × 10³ platelets/μL, 500 × 10³ platelets/μL, and 1,000 × 10³ platelets/μL). All gels contained an identical fibrinogen concentration (20 mg/mL). Fiber diameter and pore area were evaluated with scanning electron microscopy. Maximum gelation rate was assessed with spectrophotometry, and gel stiffness was determined by measuring the compressive modulus. Gel weights were measured serially over 14 days as an index of contraction (volume loss). Platelet-derived growth factor-BB and transforming growth factor-β1 concentrations were quantified with ELISAs. Fiber diameters were significantly larger and mean pore areas were significantly smaller in PRFGs than in conventional FGs. Gel weight decreased significantly over time, differed significantly between PRFGs and conventional FGs, and was significantly correlated with platelet concentration. Platelet-derived growth factor-BB and transforming growth factor-β1 concentrations were highest in gels and releasates derived from 1,000 × 10³ platelets/μL. The inclusion of platelets in FGs altered the architecture and increased the growth factor content of the resulting scaffold. Platelets may represent a useful means of modifying these gels for applications in veterinary and human regenerative medicine.

  17. Protective fibrin-sealed plication of the small bowel in recurrent laparotomy.

    Science.gov (United States)

    Holland-Cunz, S; Boelter, A V; Waag, K L

    2003-09-01

    Adhesions after recurrent abdominal operations remain extremely common and are sources of severe morbidity. Fibrin-glued plication of the small gut in a meander-like formation is supposed to guarantee a decreased risk of intestinal obstruction postoperatively. This retrospective study analyses the clinical outcome after recurrent laparotomy in children treated with bowel plication by fibrin sealant. The surgical technique of performing the fibrin-glued plication is rather simple and quick: after taking off all adhesions two to four loops of the small gut are positioned so that they lie side by side. Beginning proximal fibrin [Tissucol fibrin sealant (Baxter)] is applied between the loops; approximately 20-30 s are needed to keep the loops in position until the fibrin dries. This manoeuvre is continued until all of the small gut is fixed in one block. The gut is brought back into the abdominal cavity without loosening the loops. This fixed formation by sero-serosal adhesions or mesenterial plications is supposed to guarantee postoperative free passage. The charts of 60 children who had undergone a fibrin plication of the small bowel between 1991 and 1999 were evaluated. Additionally, questionnaires were sent to all patients, and they were invited for an examination. Sixty patients (38 boys and 22 girls) received a fibrin sealant plication because of recurrent laparotomies with heavily serosal defects or recurrent ileus because of adhesions. The youngest baby was 10 days. Since 23 patients were premature the oldest patient was 11 years old. There were no intraoperative complications attributed to the method. In the postoperative period 7/60 (12%) patients had a recurrent ileus or subileus, leading in three (5%) patients to an early relaparotomy. The fibrin-glued plication of the small bowel decreases the risk of recurrent ileus or subileus considering the high figures in the literature concerning this issue. The time-saving method is very simple and easily feasible

  18. Sinus Augmentation with Biomimetic Nanostructured Matrix: Tomographic, Radiological, Histological and Histomorphometrical Results after 6 Months in Humans

    Directory of Open Access Journals (Sweden)

    Antonio Scarano

    2017-08-01

    Full Text Available Background: Many bone substitutes have been applied for sinus regeneration procedures, such as autogenous bone, inorganic bovine bone, porous and resorbable hydroxyapatite, tricalcium phosphate, bioactive glass, and blood clots. The aim of the present study was a tomographic, histological and histomorphometrical evaluation in humans, of specimens retrieved from sinuses augmented with MgHA/collagen-based scaffolds, after a healing period of 6 months.Materials and Methods: Eleven healthy patients and a total of 15 sinuses were included in this study. The maxillary sinuses were filled with commercial MgHA/collagen-based scaffolds (RegenOss with a porous three-dimensional (3D structure (Fin-Ceramica Faenza S.p.A., Faenza, Italy. These grafts have a composite design, that replicate the organization of bone structure, obtained by a technique in which a specific hybrid organic–inorganic composite is spontaneously built by a biological mechanism. The CBCT scans were done before the procedure, after the surgical protocol (T1, and 6 months after sinus surgery (T2 for implantology. Bone specimens were stored in 10% formalin solution, embedded in a glycolmethacrylate resin and sectioned by a high-precision diamond disc. Histologic and histomorphometric analysis were carried out to evaluate the graft reabsorption and bone healing.Results: The mean volume after graft elevation, calculated for each of the 15 sinuses, was 2,906 mm3 in the immediate postoperative period (5–7 days, ranging from 2,148.8 to 3,146.4 mm3. In the late postoperative period (6 months it was 2,806.7 mm3, ranging from 2,010.9 to 3,008.9 mm3. The sinuses were completely healed and no residual MgHA/collagen-based scaffolds were visible. Osteoblasts appeared actively secreting bone matrix and marrow spaces contained moderate numbers of stromal cells and vascular network. Osteoblasts were observed actively secreting osteoid matrix. The tissues present in the samples were composed of 1.9

  19. Platelet rich fibrin - a novel acumen into regenerative endodontic therapy

    Directory of Open Access Journals (Sweden)

    Kavita Hotwani

    2014-02-01

    Full Text Available Research into regenerative dentistry has added impetus onto the field of molecular biology. It can be documented as a prototype shift in the therapeutic armamentarium for dental disease. Regenerative endodontic procedures are widely being added to the current armamentarium of pulp therapy procedures. The regenerative potential of platelets has been deliberated. A new family of platelet concentrates called the platelet rich fibrin (PRF has been recently used by several investigators and has shown application in diverse disciplines of dentistry. This paper is intended to add light on the various prospects of PRF and clinical insights to regenerative endodontic therapy.

  20. Microbiology and pathology of fibrinous pericarditis in Danish slaughter pigs

    DEFF Research Database (Denmark)

    Buttenschøn, J.; Friis, N.F.; Aalbaek, B.

    1997-01-01

    The prevalence of fibrinous pericarditis detected at slaughter in Danish slaughter pigs is approximately 0.02 %. The microbiology and pathology of this disorder was studied through 46 field cases collected at slaughter from May 1994 to August 1995. Mycoplasmas (Mycoplasma hyopneumoniae, M...... from the pericardium in three cases. The pericardium was sterile in three cases and contained a low grade mixed flora in two; the latter is believed to be a result of the slaughter procedure rather than an infection. No chlamydiae were found in connection with the microbiological examination...

  1. Fibrinous anterior uveitis following laser in situ keratomileusis

    Directory of Open Access Journals (Sweden)

    Parmar Pragya

    2009-01-01

    Full Text Available A 29-year-old woman who underwent laser in situ keratomileusis (LASIK for myopic astigmatism in both eyes presented with severe pain, photophobia and decreased visual acuity in the left eye eight days after surgery. Examination revealed severe anterior uveitis with fibrinous exudates in the anterior chamber, flap edema and epithelial bullae. Laboratory investigations for uveitis were negative and the patient required systemic and intensive topical steroids with cycloplegics to control the inflammation. This case demonstrates that severe anterior uveitis may develop after LASIK and needs prompt and vigorous management for resolution.

  2. Association Between Disruption of Fibrin Sheaths Using Percutaneous Transluminal Angioplasty Balloons and Late Onset of Central Venous Stenosis

    International Nuclear Information System (INIS)

    Ni, Nina; Mojibian, Hamid; Pollak, Jeffrey; Tal, Michael

    2011-01-01

    To compare the rates of central venous stenosis in patients undergoing hemodialysis who underwent disruption of fibrin sheath with percutaneous transluminal angioplasty balloons and those who underwent over-the-wire catheter exchange. This study is a retrospective review of 209 percutaneous transluminal angioplasty balloon disruption and 1304 over-the-wire catheter exchange procedures performed in 753 patients. Approval from the Human Investigations Committee was obtained for this study. Up to 10-year follow-up was performed. A χ 2 test was used to compare the rates of central venous stenosis after balloon disruption versus catheter exchange. A t-test was used to compare time to central venous stenosis development. Of the 753 patients in the study, 127 patients underwent balloon disruption of fibrin sheath and 626 had catheter exchange. Within the balloon disruption group, 18 (14.2%) of 127 patients subsequently developed central venous stenosis, compared with 44 (7.0%) of 626 in the catheter exchange group (P 2 test). Time to central venous stenosis development was approximately 3 years in both groups and not significantly different (1371 and 1010 days, P = 0.20). A total of 25.2% of patients in the balloon disruption group had four or more subsequent catheter exchanges, versus 12.6% in the catheter exchange group (P 2 test). In conclusions, there is a possible association between percutaneous transluminal angioplasty balloon disruption of fibrin sheath and late-onset central venous stenosis. Because venography was not routinely performed in catheter exchange patients, future randomized studies are necessary to confirm these findings.

  3. Advanced platelet-rich fibrin: a new concept for cell-based tissue engineering by means of inflammatory cells.

    Science.gov (United States)

    Ghanaati, Shahram; Booms, Patrick; Orlowska, Anna; Kubesch, Alica; Lorenz, Jonas; Rutkowski, Jim; Landes, Constantin; Sader, Robert; Kirkpatrick, Cj; Choukroun, Joseph

    2014-12-01

    Choukroun's platelet-rich fibrin (PRF) is obtained from blood without adding anticoagulants. In this study, protocols for standard platelet-rich fibrin (S-PRF) (2700 rpm, 12 minutes) and advanced platelet-rich fibrin (A-PRF) (1500 rpm, 14 minutes) were compared to establish by histological cell detection and histomorphometrical measurement of cell distribution the effects of the centrifugal force (speed and time) on the distribution of cells relevant for wound healing and tissue regeneration. Immunohistochemistry for monocytes, T and B -lymphocytes, neutrophilic granulocytes, CD34-positive stem cells, and platelets was performed on clots produced from four different human donors. Platelets were detected throughout the clot in both groups, although in the A-PRF group, more platelets were found in the distal part, away from the buffy coat (BC). T- and B-lymphocytes, stem cells, and monocytes were detected in the surroundings of the BC in both groups. Decreasing the rpm while increasing the centrifugation time in the A-PRF group gave an enhanced presence of neutrophilic granulocytes in the distal part of the clot. In the S-PRF group, neutrophils were found mostly at the red blood cell (RBC)-BC interface. Neutrophilic granulocytes contribute to monocyte differentiation into macrophages. Accordingly, a higher presence of these cells might be able to influence the differentiation of host macrophages and macrophages within the clot after implantation. Thus, A-PRF might influence bone and soft tissue regeneration, especially through the presence of monocytes/macrophages and their growth factors. The relevance and feasibility of this tissue-engineering concept have to be proven through in vivo studies.

  4. Effect of centrifugation time on growth factor and MMP release of an experimental platelet-rich fibrin-type product.

    Science.gov (United States)

    Eren, Gülnihal; Gürkan, Ali; Atmaca, Harika; Dönmez, Ayhan; Atilla, Gül

    2016-07-01

    Platelet-rich fibrin (PRF) has a controlled release of growth factors due to the fibrin matrix structure. Different centrifugation protocols were suggested for PRF preparation. Since the derivation method of PRF can alter its contents, in the present study it is aimed to investigate the cell contents and transforming growth factor beta-1 (TGF-β1), platelet-derived growth factor (PDGF-AB), vascular endothelial growth factor (VEGF), matrix metalloproteinase (MMP)-1 and-8 release from experimental PRF-type membranes obtained with different centrifugation times at 400 gravity. Three blood samples were collected from 20 healthy non-smoker volunteers. One tube was used for whole blood analyses. The other two tubes were centrifuged at 400 g for 10 minutes (group A) or 12 minutes (group B). Each experimental PRF-type membrane was placed in Dulbecco's Modified Eagle's Medium (DMEM)and at 1, 24 and 72 hours, TGF-β1, PDGF-AB, VEGF, MMP-1 and -8 release amounts were analysed by enzyme-linked immunosorbent assay (ELISA). The blood cell count of membranes was determined by subtracting plasma supernatant and red blood cell (RBC) mixture from the whole blood cell counts. At 72 hours, the VEGF level of group B was statistically higher than that of group A (p = 0.040). The centrifugation time was not found to influence the release of other growth factors, enzymes and cell counts. Within the limits of the present study, it might be suggested that centrifugation time at a constant gravity has a significant effect on the VEGF levels released from experimental PRF-type membrane. It can be concluded that due to the importance of VEGF in the tissue healing process, membranes obtained at 12-minute centrifugation time may show a superior potential in wound healing.

  5. Use of additives, scaffolds and extracellular matrix components for improvement of human pancreatic islet outcomes in vitro: A systematic review.

    Science.gov (United States)

    Lemos, Natália Emerim; de Almeida Brondani, Letícia; Dieter, Cristine; Rheinheimer, Jakeline; Bouças, Ana Paula; Bauermann Leitão, Cristiane; Crispim, Daisy; Bauer, Andrea Carla

    2017-09-03

    Pancreatic islet transplantation is an established treatment to restore insulin independence in type 1 diabetic patients. Its success rates have increased lately based on improvements in immunosuppressive therapies and on islet isolation and culture. It is known that the quality and quantity of viable transplanted islets are crucial for the achievement of insulin independence and some studies have shown that a significant number of islets are lost during culture time. Thus, in an effort to improve islet yield during culture period, researchers have tested a variety of additives in culture media as well as alternative culture devices, such as scaffolds. However, due to the use of different categories of additives or devices, it is difficult to draw a conclusion on the benefits of these strategies. Therefore, the aim of this systematic review was to summarize the results of studies that described the use of medium additives, scaffolds or extracellular matrix (ECM) components during human pancreatic islets culture. PubMed and Embase repositories were searched. Of 5083 articles retrieved, a total of 37 articles fulfilled the eligibility criteria and were included in the review. After data extraction, articles were grouped as follows: 1) "antiapoptotic/anti-inflammatory/antioxidant," 2) "hormone," 3) "sulphonylureas," 4) "serum supplements," and 5) "scaffolds or ECM components." The effects of the reviewed additives, ECM or scaffolds on islet viability, apoptosis and function (glucose-stimulated insulin secretion - GSIS) were heterogeneous, making any major conclusion hard to sustain. Overall, some "antiapoptotic/anti-inflammatory/antioxidant" additives decreased apoptosis and improved GSIS. Moreover, islet culture with ECM components or scaffolds increased GSIS. More studies are needed to define the real impact of these strategies in improving islet transplantation outcomes.

  6. In vivo near infrared (NIRS) sensor attachment using fibrin bioadhesive

    Science.gov (United States)

    Macnab, Andrew; Pagano, Roberto; Kwon, Brian; Dumont, Guy; Shadgan, Babak

    2018-02-01

    Background: `Tisseel' (Baxter Healthcare, Deerfield, IL) is a fibrin-based sealant that is commonly used during spine surgery to augment dural repairs. We wish to intra-operatively secure a near infrared spectroscopy (NIRS) sensor to the dura in order to monitor the tissue hemodynamics of the underlying spinal cord. To determine if `Tisseel' sealant adversely attenuates NIR photon transmission. Methods: We investigated `Tisseel' in both an in vitro and in vivo paradigm. For in vitro testing, we used a 1 mm pathlength cuvette containing either air or `Tisseel' interposed between a NIR light source (760 and 850 nm) and a photodiode detector and compared transmittance. For in vivo testing, a continuous wave (760 and 850 nm) spatiallyresolved NIRS device was placed over the triceps muscle using either conventional skin apposition (overlying adhesive bandage) or bioadhesion with `Tisseel'. Raw optical data and tissue saturation index (TSI%) collected at rest were compared. Results: In-vitro NIR light absorption by `Tisseel' was very high, with transmittance reduced by 95% compared to air. In-vivo muscle TSI% values were 80% with conventional attachment and 20% using fibrin glue. Conclusion: The optical properties of `Tisseel' significantly attenuate NIR light during in-vitro transmittance and critically compromise photon transmission in-vivo.

  7. Study of some parameters of the fibrinogen - fibrin transformation reaction

    International Nuclear Information System (INIS)

    Hollard, D.; Suscillon, M.; Marcille, G.; Rambaud, F.; Baloyan, M.

    1966-01-01

    The authors studied the action of some parameters on the reaction of transformation fibrinogen-fibrin. The five parameters studied are: the concentration of substratum: a certain quantity of enzyme determines an optimum quantity of fibrinogen; the concentration of enzyme: a certain quantity of substratum defines an optimum quantity of enzyme, beyond which the excess of enzyme is unable to act, the substratum being saturated by the enzyme; the concentration of Ca ions: between 0,07 and 0,10 mg of Ca by mg of fibrinogen, the reaction appears with a great speed. Between 0,02 and 0,40 mg of Ca by mg of fibrinogen the fibrin stabilisation is possible, the FSF can act only inside the definite bounds; the ph of the solution: the reaction of the transformation appears with its maximum intensity on physiological ph, the polymerisation is not possible on acid ph; the temperature has an effect which could not really be verified owing to the fact that the technical realisation is difficult. (author) [fr

  8. Hyaline Articular Matrix Formed by Dynamic Self-Regenerating Cartilage and Hydrogels.

    Science.gov (United States)

    Meppelink, Amanda M; Zhao, Xing; Griffin, Darvin J; Erali, Richard; Gill, Thomas J; Bonassar, Lawrence J; Redmond, Robert W; Randolph, Mark A

    2016-07-01

    Injuries to the articular cartilage surface are challenging to repair because cartilage possesses a limited capacity for self-repair. The outcomes of current clinical procedures aimed to address these injuries are inconsistent and unsatisfactory. We have developed a novel method for generating hyaline articular cartilage to improve the outcome of joint surface repair. A suspension of 10(7) swine chondrocytes was cultured under reciprocating motion for 14 days. The resulting dynamic self-regenerating cartilage (dSRC) was placed in a cartilage ring and capped with fibrin and collagen gel. A control group consisted of chondrocytes encapsulated in fibrin gel. Constructs were implanted subcutaneously in nude mice and harvested after 6 weeks. Gross, histological, immunohistochemical, biochemical, and biomechanical analyses were performed. In swine patellar groove, dSRC was implanted into osteochondral defects capped with collagen gel and compared to defects filled with osteochondral plugs, collagen gel, or left empty after 6 weeks. In mice, the fibrin- and collagen-capped dSRC constructs showed enhanced contiguous cartilage matrix formation over the control of cells encapsulated in fibrin gel. Biochemically, the fibrin and collagen gel dSRC groups were statistically improved in glycosaminoglycan and hydroxyproline content compared to the control. There was no statistical difference in the biomechanical data between the dSRC groups and the control. The swine model also showed contiguous cartilage matrix in the dSRC group but not in the collagen gel and empty defects. These data demonstrate the survivability and successful matrix formation of dSRC under the mechanical forces experienced by normal hyaline cartilage in the knee joint. The results from this study demonstrate that dSRC capped with hydrogels successfully engineers contiguous articular cartilage matrix in both nonload-bearing and load-bearing environments.

  9. Nonuniform Internal Structure of Fibrin Fibers: Protein Density and Bond Density Strongly Decrease with Increasing Diameter

    Directory of Open Access Journals (Sweden)

    Wei Li

    2017-01-01

    Full Text Available The major structural component of a blood clot is a meshwork of fibrin fibers. It has long been thought that the internal structure of fibrin fibers is homogeneous; that is, the protein density and the bond density between protofibrils are uniform and do not depend on fiber diameter. We performed experiments to investigate the internal structure of fibrin fibers. We formed fibrin fibers with fluorescently labeled fibrinogen and determined the light intensity of a fiber, I, as a function of fiber diameter, D. The intensity and, thus, the total number of fibrin molecules in a cross-section scaled as D1.4. This means that the protein density (fibrin per cross-sectional area, ρp, is not homogeneous but instead strongly decreases with fiber diameter as D-0.6. Thinner fibers are denser than thicker fibers. We also determined Young’s modulus, Y, as a function of fiber diameter. Y decreased strongly with increasing D; Y scaled as D-1.5. This implies that the bond density, ρb, also scales as D-1.5. Thinner fibers are stiffer than thicker fibers. Our data suggest that fibrin fibers have a dense, well-connected core and a sparse, loosely connected periphery. In contrast, electrospun fibrinogen fibers, used as a control, have a homogeneous cross-section.

  10. Atherosclerotic arterial remodeling and the localization of macrophages and matrix metalloproteases 1, 2 and 9 in the human coronary artery

    NARCIS (Netherlands)

    Pasterkamp, G.; Schoneveld, A. H.; Hijnen, D. J.; de Kleijn, D. P.; Teepen, H.; van der Wal, A. C.; Borst, C.

    2000-01-01

    Atherosclerotic luminal narrowing is determined by plaque mass and the mode of geometrical remodeling. Recently, we reported that the type of atherosclerotic remodeling is associated with the presence of histological markers for plaque vulnerability. Inflammation and matrix degrading proteases

  11. The disulfide isomerase ERp57 is required for fibrin deposition in vivo.

    Science.gov (United States)

    Zhou, J; Wu, Y; Wang, L; Rauova, L; Hayes, V M; Poncz, M; Essex, D W

    2014-11-01

    ERp57 is required for platelet function; however, whether ERp57 contributes to fibrin generation is unknown. Using an inhibitory anti-ERp57 antibody (mAb1), Pf4-Cre/ERp57(fl/fl) mice, Tie2-Cre/ERp57(fl/fl) mice, and mutants of ERp57, we analyzed the function of ERp57 in laser-induced thrombosis. Fibrin deposition was decreased in Pf4-Cre/ERp57(fl/fl) mice, consistent with a role for platelet ERp57 in fibrin generation. Fibrin deposition was further decreased with infusion of mAb1 and in Tie2-Cre/ERp57(fl/fl) mice, consistent with endothelial cells also contributing to fibrin deposition. Infusion of eptibifatide inhibited platelet and fibrin deposition, confirming a role for platelets in fibrin deposition. Infusion of recombinant ERp57 corrected the defect in fibrin deposition but not platelet accumulation, suggesting a direct effect of ERp57 on coagulation. mAb1 inhibited thrombin generation in vitro, consistent with a requirement for ERp57 in coagulation. Platelet accumulation was decreased to similar extents in Pf4-Cre/ERp57(fl/fl) mice, Tie2-Cre/ERp57(fl/fl) mice and normal mice infused with mAb1. Infusion of completely inactivated ERp57 or ERp57 with a non-functional second active site inhibited fibrin deposition and platelet accumulation, indicating that the isomerase activity of the second active site is required for these processes. ERp57 regulates thrombosis via multiple targets. © 2014 International Society on Thrombosis and Haemostasis.

  12. Platelet glycoprotein VI binds to polymerized fibrin and promotes thrombin generation.

    Science.gov (United States)

    Mammadova-Bach, Elmina; Ollivier, Véronique; Loyau, Stéphane; Schaff, Mathieu; Dumont, Bénédicte; Favier, Rémi; Freyburger, Geneviève; Latger-Cannard, Véronique; Nieswandt, Bernhard; Gachet, Christian; Mangin, Pierre H; Jandrot-Perrus, Martine

    2015-07-30

    Fibrin, the coagulation end product, consolidates the platelet plug at sites of vascular injury and supports the recruitment of circulating platelets. In addition to integrin αIIbβ3, another as-yet-unidentified receptor is thought to mediate platelet interaction with fibrin. Platelet glycoprotein VI (GPVI) interacts with collagen and several other adhesive macromolecules. We evaluated the hypothesis that GPVI could be a functional platelet receptor for fibrin. Calibrated thrombin assays using platelet-rich plasma (PRP) showed that tissue factor-triggered thrombin generation was impaired in GPVI-deficient patients and reduced by the anti-GPVI Fab 9O12. Assays on reconstituted PRP and PRP from fibrinogen-deficient patients revealed a fibrinogen-dependent enhancement of thrombin generation, which relied on functional GPVI. The effect of GPVI was found to depend on fibrin polymerization. A binding assay showed a specific interaction between GPVI-Fc and fibrin, inhibited by the Fab 9O12. This Fab also reduced platelet adhesion to fibrin at low (300 s(-1)) and high (1500 s(-1)) wall shear rates. Platelets adherent to fibrin displayed shape change, exposure of procoagulant phospholipids, and the formation of small clots. When hirudinated blood was perfused at 1500 s(-1) over preformed fibrin-rich clots, the Fab 9O12 decreased the recruitment of platelets by up to 85%. This study identifies GPVI as a platelet receptor for polymerized fibrin with 2 major functions: (1) amplification of thrombin generation and (2) recruitment of circulating platelets to clots. These so-far-unrecognized properties of GPVI confer on it a key role in thrombus growth and stabilization. © 2015 by The American Society of Hematology.

  13. Effects of fibrin, thrombin, and blood on breast capsule formation in a preclinical model.

    Science.gov (United States)

    Marques, Marisa; Brown, Spencer A; Cordeiro, Natália D S; Rodrigues-Pereira, Pedro; Cobrado, M Luís; Morales-Helguera, Aliuska; Lima, Nuno; Luís, André; Mendanha, Mário; Gonçalves-Rodrigues, Acácio; Amarante, José

    2011-03-01

    The root cause of capsular contracture (CC) associated with breast implants is unknown. Recent evidence points to the possible role of fibrin and bacteria in CC formation. The authors sought to determine whether fibrin, thrombin, and blood modulated the histological and microbiological outcomes of breast implant capsule formation in a rabbit model. The authors carried out a case-control study to assess the influence of fibrin, thrombin, and blood on capsule wound healing in a rabbit model. Eighteen New Zealand white rabbits received four tissue expanders. One expander acted as a control, whereas the other expander pockets received one of the following: fibrin glue, rabbit blood, or thrombin sealant. Intracapsular pressure/volume curves were compared among the groups, and histological and microbiological evaluations were performed (capsules, tissue expanders, rabbit skin, and air). The rabbits were euthanized at two or four weeks. At four weeks, the fibrin and thrombin expanders demonstrated significantly decreased intracapsular pressure compared to the control group. In the control and fibrin groups, mixed inflammation correlated with decreased intracapsular pressure, whereas mononuclear inflammation correlated with increased intracapsular pressure. The predominant isolate in the capsules, tissue expanders, and rabbit skin was coagulase-negative staphylococci. For fibrin and thrombin, both cultures that showed an organism other than staphylococci and cultures that were negative were associated with decreased intracapsular pressure, whereas cultures positive for staphylococci were associated with increased intracapsular pressure. Fibrin application during breast implantation may reduce rates of CC, but the presence of staphylococci is associated with increased capsule pressure even in the presence of fibrin, so care should be taken to avoid bacterial contamination.

  14. Tumor necrosis factor-α-accelerated degradation of type I collagen in human skin is associated with elevated matrix metalloproteinase (MMP)-1 and MMP-3 ex vivo

    DEFF Research Database (Denmark)

    Ågren, Magnus S; Schnabel, Reinhild; Christensen, Lise H

    2015-01-01

    Tumor necrosis factor (TNF)-α induces matrix metalloproteinases (MMPs) that may disrupt skin integrity. We have investigated the effects and mechanisms of exogenous TNF-α on collagen degradation by incubating human skin explants in defined serum-free media with or without TNF-α (10ng/ml) in the a......Tumor necrosis factor (TNF)-α induces matrix metalloproteinases (MMPs) that may disrupt skin integrity. We have investigated the effects and mechanisms of exogenous TNF-α on collagen degradation by incubating human skin explants in defined serum-free media with or without TNF-α (10ng...... tissue-derived collagenolytic activity with TNF-α exposure was blocked by neutralizing MMP-1 monoclonal antibody and was not due to down-regulation of tissue inhibitor of metalloproteinase-1. TNF-α increased production (pendogenous MMP-1...

  15. Matrix theory

    CERN Document Server

    Franklin, Joel N

    2003-01-01

    Mathematically rigorous introduction covers vector and matrix norms, the condition-number of a matrix, positive and irreducible matrices, much more. Only elementary algebra and calculus required. Includes problem-solving exercises. 1968 edition.

  16. Fibrin structural and diffusional analysis suggests that fibers are permeable to solute transport.

    Science.gov (United States)

    Leonidakis, Kimon Alexandros; Bhattacharya, Pinaki; Patterson, Jennifer; Vos, Bart E; Koenderink, Gijsje H; Vermant, Jan; Lambrechts, Dennis; Roeffaers, Maarten; Van Oosterwyck, Hans

    2017-01-01

    Fibrin hydrogels are promising carrier materials in tissue engineering. They are biocompatible and easy to prepare, they can bind growth factors and they can be prepared from a patient's own blood. While fibrin structure and mechanics have been extensively studied, not much is known about the relation between structure and diffusivity of solutes within the network. This is particularly relevant for solutes with a size similar to that of growth factors. A novel methodological approach has been used in this study to retrieve quantitative structural characteristics of fibrin hydrogels, by combining two complementary techniques, namely confocal fluorescence microscopy with a fiber extraction algorithm and turbidity measurements. Bulk rheological measurements were conducted to determine the impact of fibrin hydrogel structure on mechanical properties. From these measurements it can be concluded that variations in the fibrin hydrogel structure have a large impact on the rheological response of the hydrogels (up to two orders of magnitude difference in storage modulus) but only a moderate influence on the diffusivity of dextran solutes (up to 25% difference). By analyzing the diffusivity measurements by means of the Ogston diffusion model we further provide evidence that individual fibrin fibers can be semi-permeable to solute transport, depending on the average distance between individual protofibrils. This can be important for reducing mass transport limitations, for modulating fibrinolysis and for growth factor binding, which are all relevant for tissue engineering. Fibrin is a natural biopolymer that has drawn much interest as a biomimetic carrier in tissue engineering applications. We hereby use a novel combined approach for the structural characterization of fibrin networks based on optical microscopy and light scattering methods that can also be applied to other fibrillar hydrogels, like collagen. Furthermore, our findings on the relation between solute transport

  17. The provisional matrix: setting the stage for tissue repair outcomes.

    Science.gov (United States)

    Barker, Thomas H; Engler, Adam J

    2017-07-01

    Since its conceptualization in the 1980s, the provisional matrix has often been characterized as a simple fibrin-containing scaffold for wound healing that supports the nascent blood clot and is functionally distinct from the basement membrane. However subsequent advances have shown that this matrix is far from passive, with distinct compositional differences as the wound matures, and providing an active role for wound remodeling. Here we review the stages of this matrix, provide an update on the state of our understanding of provisional matrix, and present some of the outstanding issues related to the provisional matrix, its components, and their assembly and use in vivo. Copyright © 2017. Published by Elsevier B.V.

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

    Directory of Open Access Journals (Sweden)

    Olga Tatti

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

  19. Platelet-rich fibrin: a boon in regenerative endodontics.

    Science.gov (United States)

    Rebentish, Priyanka D; Umashetty, Girish; Kaur, Harpreet; Doizode, Trupthi; Kaslekar, Mithun; Chowdhury, Shouvik

    2016-12-01

    Research into regenerative dentistry has contributed momentum to the field of molecular biology. Periapical surgery aims at removing periapical pathology to achieve complete wound healing and regeneration of bone and periodontal tissue. Regenerative endodontic procedures are widely being added to the current armamentarium of pulp therapy procedures. The regenerative potential of platelets has been deliberated. Platelet-rich fibrin (PRF) is a wonderful tissue-engineering product and has recently gained much popularity due its promising results in wound healing bone induction. The features of this product are an attribute of platelets which, after cellular interactions, release growth factors and have shown application in diverse disciplines of dentistry. This paper is intended to shed light onto the various prospects of PRF and to provide clinical insight into regenerative endodontic therapy.

  20. Platelet-rich-fibrin: A novel root coverage approach

    Directory of Open Access Journals (Sweden)

    Anilkumar K

    2009-01-01

    Full Text Available Treatment of gingival recession has become an important therapeutic issue due to increasing cosmetic demand. Multiple surgical procedures have been developed to obtain predictable esthetic root coverage. More specifically, after periodontal regenerative surgery, the aim is to achieve complete wound healing and regeneration of the periodontal unit. A recent innovation in dentistry is the preparation and use of platelet-rich plasma (PRP, a concentrated suspension of the growth factors, found in platelets. These growth factors are involved in wound healing and postulated as promoters of tissue regeneration. This paper reports the use of PRF membrane for root coverage on the labial surfaces of the mandibular anterior teeth. This was accomplished using laterally displaced flap technique with platelet rich fibrin (PRF membrane at the recipient site.

  1. Fibrin Glue Does Not Improve Healing of Gastrointestinal Anastomoses

    DEFF Research Database (Denmark)

    Nordentoft, Tyge; Pommergaard, Hans-Christian; Rosenberg, Jacob

    2015-01-01

    BACKGROUND/AIM: Anastomotic leakage remains a frequent and serious complication in gastrointestinal surgery. In order to reduce its incidence, several clinical and experimental studies on anastomotic sealing have been performed. In a number of these studies, the sealing material has been fibrin...... glue (FG), and the results in individual studies have been varying. The positive effect of anastomotic sealing with FG might be due to the mechanical/physical properties, the increased healing of the anastomoses or both. The aim of this systematic review was to evaluate the existing evidence....... It is consequently plausible that the positive effect of FG sealing of gastrointestinal anastomoses, if there is any, may be due to a mechanical sealing effect rather than due to improved healing per se....

  2. Co-transfection of decorin and interleukin-10 modulates pro-fibrotic extracellular matrix gene expression in human tenocyte culture

    Science.gov (United States)

    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.

  3. Platelet-rich fibrin or platelet-rich plasma – which one is better? an opinion

    Directory of Open Access Journals (Sweden)

    Shweta Bansal

    2017-01-01

    Full Text Available The healing of hard and soft tissue in mediated by a wide range of intracellular and extracellular events that are regulated by signaling proteins. Platelets can play a crucial role in periodontal regeneration as they are the reservoirs of growth factors and cytokines which are the key factors for regeneration of bone and maturation of soft tissue. Platelet-rich plasma (PRP is first generation platelet concentrate. However, the short duration of cytokine release and its poor mechanical properties have resulted in search of new material. Platelet-rich fibrin (PRF is a natural fibrin-based biomaterial prepared from an anticoagulant-free blood harvest without any artificial biochemical modification (no bovine thrombin is required that allows obtaining fibrin membranes enriched with platelets and growth factors. The slow polymerization during centrifugation, fibrin-based structure, ease of preparation, minimal expense makes PRF somewhat superior in some aspect to PRP.

  4. Thyroid Dysfunction and Fibrin Network Structure: A Mechanism for Increased Thrombotic Risk in Hyperthyroid Individuals

    NARCIS (Netherlands)

    Hooper, J. M. W.; Stuijver, D. J. F.; Orme, S. M.; van Zaane, B.; Hess, K.; Gerdes, V. E.; Phoenix, F.; Rice, P.; Smith, K. A.; Alzahrani, S. H.; Standeven, K. F.; Ajjan, R. A.

    2012-01-01

    Context: Hyperthyroidism is associated with increased thrombosis risk, and fibrin clot structure determines susceptibility to vascular thrombotic events. Objective: Our objective was to investigate clot formation and lysis in hyperthyroidism using observational and interventional studies. Design: Ex

  5. Sealing of Gastrointestinal Anastomoses with a Fibrin Glue-Coated Collagen Patch: A Safety Study

    DEFF Research Database (Denmark)

    Nordentoft, Tyge; Rømer, John; Sørensen, Michael

    2007-01-01

    gastrointestinal anastomoses with a collagen patch coated with fibrin glue. The study is a prospective, experimental animal study comparing sealed and unsealed gastrointestinal anastomoses. Laparotomy was performed in 11 pigs under general anesthesia. In each pig two anastomoses were performed on the small......Sealing of anastomoses has previously been tested with several methods, including sealing with liquid fibrin glue. Sealing with a collagen patch coated with fibrin glue components has never been systematically examined. The aim of the present study was to determine the safety of sealing...... intestine. One of the anastomoses was sealed with a collagen patch coated with fibrin glue components (TachoSil). The other anastomosis contained no sealing. The pigs were observed for 1 to 6 weeks. The observation period was followed by in vivo examination under general anesthesia and included observation...

  6. Effect of Titanium-prepared Platelet-rich Fibrin Treatment on the ...

    African Journals Online (AJOL)

    2018-02-07

    Feb 7, 2018 ... Defects of Patients with Chronic Periodontitis: A Randomized. Controlled ... Faculty of Dentistry Department of Periodontology, Kirikkale. University ..... using leukocyte and platelet-rich fibrin: A report of a case. Case. Rep Dent ...

  7. Effect of Titanium-prepared Platelet-rich Fibrin Treatment on the ...

    African Journals Online (AJOL)

    Effect of Titanium-prepared Platelet-rich Fibrin Treatment on the Angiogenic Biomarkers in Gingival Crevicular Fluid in Infrabony Defects of Patients with Chronic Periodontitis: A Randomized Controlled Clinical Trial.

  8. The use of snake venom derived fibrin glue in hysterorrhaphy of ovine caesarean surgery

    OpenAIRE

    CHALHOUB, M.; PRESTES, N. C.; LOPES, M. D.; ROCHA, N. S.; THOMAZINI-SANTOS, I. A.; MENDES-GIANNINI, M.J.

    2000-01-01

    Fibrin glue has been used on its own or in conjunction with suturing materials to promote hemostasis, reduce adherence, strengthen the wound site, and improve healing. Snake venom derived fibrin glue was evaluated as an alternative to conventional uterine suturing after ovine caesarean surgery. Twenty-eight pregnant ewes of known mating date were used. The animals submitted to conventional caesarean sections showed a better wound healing process. As expected, all the operated animals had reta...

  9. Fibrin Clots Are Equilibrium Polymers That Can Be Remodeled Without Proteolytic Digestion

    OpenAIRE

    Chernysh, Irina N.; Nagaswami, Chandrasekaran; Purohit, Prashant K.; Weisel, John W.

    2012-01-01

    Fibrin polymerization is a necessary part of hemostasis but clots can obstruct blood vessels and cause heart attacks and strokes. The polymerization reactions are specific and controlled, involving strong knob-into-hole interactions to convert soluble fibrinogen into insoluble fibrin. It has long been assumed that clots and thrombi are stable structures until proteolytic digestion. On the contrary, using the technique of fluorescence recovery after photobleaching, we demonstrate here that the...

  10. Does Fibrin Sealant Reduce Seroma after Immediate Breast Reconstruction Utilizing a Latissimus Dorsi Myocutaneous Flap?

    Directory of Open Access Journals (Sweden)

    Han Gyu Cha

    2012-09-01

    Full Text Available Background The most common complication of latissimus dorsi myocutaneous flap in breastreconstruction is seroma formation in the back. Many clinical studies have shown that fibrinsealant reduces seroma formation. We investigated any statistically significant differences inpostoperative drainage and seroma formation when utilizing the fibrin sealant on the site ofthe latissimus dorsi myocutaneous flap harvested for immediate breast reconstruction afterskin-sparing partial mastectomy.Methods A total of 46 patients underwent immediate breast reconstruction utilizing alatissimus dorsi myocutaneous island flap. Of those, 23 patients underwent the procedurewithout fibrin sealant and the other 23 were administered the fibrin sealant. All flaps wereelevated with manual dissection by the same surgeon and were analyzed to evaluate thepotential benefits of the fibrin sealant. The correlation analysis and Mann-Whitney U testwere used for analyzing the drainage volume according to age, weight of the breast specimen,and body mass index.Results Although not statistically significant, the cumulative drainage fluid volume was higherin the control group until postoperative day 2 (530.1 mL compared to 502.3 mL, but thefibrin sealant group showed more drainage beginning on postoperative day 3. The donor sitecomparisons showed the fibrin sealant group had more drainage beginning on postoperativeday 3 and the drain was removed 1 day earlier in the control group.Conclusions The use of fibrin sealant resulted in no reduction of seroma formation. Becausethe benefits of the fibrin sealant are not clear, the use of fibrin sealant must be fully discussedwith patients before its use as a part of informed consent.

  11. Tunneled dialysis catheter exchange with fibrin sheath disruption is not associated with increased rate of bacteremia.

    Science.gov (United States)

    Valliant, Amanda M; Chaudhry, Muhammad K; Yevzlin, Alexander S; Astor, Brad; Chan, Micah R

    2015-01-01

    Tunneled dialysis catheters are the most common form of vascular access among incident dialysis patients in the United States. Fibrin sheath formation is a frequent cause of late catheter dysfunction requiring an exchange procedure with balloon disruption of the fibrin sheath. It is unknown whether fibrin sheath disruption is associated with increased incidence of bacteremia or catheter failure. We reviewed all tunneled dialysis catheter exchange procedures at the University of Wisconsin between January 2008 and December 2011. The primary outcome was incidence of bacteremia, defined as positive blood cultures within 2 weeks of the procedure. Catheter failure, requiring intervention or replacement, was examined as a secondary outcome. Baseline characteristics examined included diabetic status, gender, race and age. A total of 163 procedures were reviewed; 67 (41.1%) had fibrin sheath disruption and 96 did not. Bacteremia occurred in 4.5% (3/67) of those with and 3.1% (3/97) of those without fibrin sheath disruption (p=0.65). Fibrin sheath disruption was not significantly associated with the risk of catheter failure (adjusted hazard ratio [aHR]=1.34; 95% confidence interval [CI]: 0.87-2.10; p=0.18). Diabetes was associated with greater risk of catheter failure (aHR=1.88; 95% CI: 1.19-2.95; p=0.006), whereas higher age was associated with a lower risk of catheter failure (aHR per 10 years=0.83; 95% CI: 0.72-0.96; p=0.01). This study demonstrates that there is no significant increase in bacteremia and subsequent catheter dysfunction rates after fibrin sheath disruption compared to simple over the wire exchange. These results are encouraging given the large numbers of patients utilizing tunneled catheters for initial hemodialysis access and the known rates of fibrin sheath formation leading to catheter failure.

  12. Effects of transforming growth factor-beta1 on cell motility, collagen gel contraction, myofibroblastic differentiation, and extracellular matrix expression of human adipose-derived stem cell.

    Science.gov (United States)

    Kakudo, Natsuko; Kushida, Satoshi; Suzuki, Kenji; Ogura, Tsunetaka; Notodihardjo, Priscilla Valentin; Hara, Tomoya; Kusumoto, Kenji

    2012-12-01

    Human adipose-derived stem cells (ASCs) are adult pluripotent stem cells, and their usefulness in plastic surgery has garnered attention in recent years. Although, there have been expectations that ASCs might function in wound repair and regeneration, no studies to date have examined the role of ASCs in the mechanism that promotes wound-healing. Transforming growth factor-beta1 (TGF-β1) is a strong candidate cytokine for the triggering of mesenchymal stem cell migration, construction of extracellular matrices, and differentiation of ASCs into myofibroblasts. Cell proliferation, motility, and differentiation, as well as extracellular matrix production, play an important role in wound-healing. We have evaluated the capacity of ASCs to proliferate and their potential to differentiate into phenotypic myofibroblasts, as well as their cell motility and collagen gel contraction ability, when cultured with TGF-β1. Cell motility was analyzed using a wound-healing assay. ASCs that differentiated into myofibroblasts expressed the gene for alpha-smooth muscle actin, and its protein expression was detected immunohistochemically. The extracellular matrix expression in ASCs was evaluated using real-time RT-PCR. Based on the results, we conclude that human ASCs have the potential for cell motility, extracellular matrix gene expression, gel contraction, and differentiation into myofibroblasts and, therefore, may play an important role in the wound-healing process.

  13. Evaluation of gold nanotracers to track adipose-derived stem cells in a PEGylated fibrin gel for dermal tissue engineering applications

    Directory of Open Access Journals (Sweden)

    Chung E

    2013-01-01

    Full Text Available Eunna Chung,1 Seung Yun Nam,1,2 Laura M Ricles,1 Stanislav Y Emelianov,1,2 Laura J Suggs11Department of Biomedical Engineering, 2Department of Electrical and Computer Engineering, The University of Texas at Austin, Austin, TX, USAAbstract: Evaluating the regenerative capacity of a tissue-engineered device in a noninvasive and synchronous manner is critical to determining the mechanisms for success in clinical applications. In particular, directly tracking implanted cells in a three-dimensional (3D scaffold is desirable in that it enables the monitoring of cellular activity in a specific and localized manner. The authors' group has previously demonstrated that the PEGylation of fibrin results in a 3D scaffold that supports morphologic and phenotypic changes in mesenchymal stem cells that may be advantageous in wound healing applications. Recently, the authors have evaluated adipose-derived stem cells (ASCs as a mesenchymal cell source to regenerate skin and blood vessels due to their potential for proliferation, differentiation, and production of growth factors. However, tracking and monitoring ASCs in a 3D scaffold, such as a PEGylated fibrin gel, have not yet been fully investigated. In the current paper, nanoscale gold spheres (20 nm as cell tracers for ASCs cultured in a PEGylated fibrin gel were evaluated. An advanced dual-imaging modality combining ultrasound and photoacoustic imaging was utilized to monitor rat ASCs over time. The ASCs took up gold nanotracers and could be detected up to day 16 with high sensitivity using photoacoustic imaging. There were no detrimental effects on ASC morphology, network formation, proliferation, and protein expression/secretion (ie, smooth muscle α-actin, vascular endothelial growth factor, matrix metalloproteinase-2, and matrix metalloproteinase-9 associated with gold nanotracers. Therefore, utilization of gold nanotracers can be an effective strategy to monitor the regenerative process of a stem cell

  14. Analysis of the distribution of DNA repair patches in the DNA-nuclear matrix complex from human cells

    International Nuclear Information System (INIS)

    Mullenders, L.H.F.

    1983-01-01

    The distribution of ultraviolet-induced repair patches along DNA loops attached to the nuclear matrix, was investigated by digestion with DNA-degrading enzymes and neutral sucrose gradient centrifugation. When DNA was gradually removed by DNAase 1, pulse label incorporated by ultraviolet-irradiated cells during 10 min in the presence of hydroxyurea or hydroxyurea/arabinosylcytosine showed similar degradation kinetics as prelabelled DNA. No preferential association of pulse label with the nuclear matrix was observed, neither within 30 min nor 13 h after iiradiation. When the pulse label was incorporated by replicative synthesis under the same conditions, a preferential association of newly-synthesized DNA with the nuclear matrix was observed. Single-strand specific digestion with nuclease S 1 of nuclear lysates from ultraviolet-irradiated cells, pulse labelled in the presence of hydroxyurea/arabinosylcytosine, caused a release of about 70% of the prelabelled DNA and 90% of the pulse-labelled DNA from the rapidly sedimenting material in sucrose gradients. The results suggest no specific involvement of the nuclear matrix in repair synthesis, a random distribution of repair patches along the DNA loops, and simultaneously multiple incision events per DNA loop. (Auth.)

  15. Analysis of the distribution of DNA repair patches in the DNA-nuclear matrix complex from human cells

    Energy Technology Data Exchange (ETDEWEB)

    Mullenders, L.H.F. (Rijksuniversiteit Leiden (Netherlands). Lab. voor Stralengenetica en Chemische Mutagenese); Zeeland, A.A. van; Natarajan, A.T. (Cohen (J.A.) Inst. voor Radiopathologie en Stralenbescherming, Leiden (Netherlands))

    1983-09-09

    The distribution of ultraviolet-induced repair patches along DNA loops attached to the nuclear matrix, was investigated by digestion with DNA-degrading enzymes and neutral sucrose gradient centrifugation. When DNA was gradually removed by DNAase 1, pulse label incorporated by ultraviolet-irradiated cells during 10 min in the presence of hydroxyurea or hydroxyurea/arabinosylcytosine showed similar degradation kinetics as prelabelled DNA. No preferential association of pulse label with the nuclear matrix was observed, neither within 30 min nor 13 h after irradiation. When the pulse label was incorporated by replicative synthesis under the same conditions, a preferential association of newly-synthesized DNA with the nuclear matrix was observed. Single-strand specific digestion with nuclease S/sub 1/ of nuclear lysates from ultraviolet-irradiated cells, pulse labelled in the presence of hydroxyurea/arabinosylcytosine, caused a release of about 70% of the prelabelled DNA and 90% of the pulse-labelled DNA from the rapidly sedimenting material in sucrose gradients. The results suggest no specific involvement of the nuclear matrix in repair synthesis, a random distribution of repair patches along the DNA loops, and simultaneously multiple incision events per DNA loop.

  16. Synthetic inhibitors of matrix metalloproteinases prevent sulfur mustard-induced epidermal-dermal separation in human skin pieces

    NARCIS (Netherlands)

    Mol, M.A.E.; Alblas, S.W.; Hammer, A.; Benschop, H.P.

    2000-01-01

    Degradation of proteins of the basement membrane zone (BMZ) in the skin depends on the activity of proteolytic enzymes, particularly those belonging to the group of matrix metalloproteinases (MMPs). In the present study we have investigated the contribution of these enzymes to the epidermal-dermal

  17. Extracellular matrix components supporting human islet function in alginate-based immunoprotective microcapsules for treatment of diabetes

    NARCIS (Netherlands)

    Llacua Carrasco, Luis; de Haan, Bart J; Smink, Sandra A; de Vos, Paul

    In the pancreas, extracellular matrix (ECM) components play an import role in providing mechanical and physiological support, and also contribute to the function of islets. These ECM-connections are damaged during islet-isolation from the pancreas and are not fully recovered after encapsulation and

  18. Distribution of cytoskeletal proteins, integrins, leukocyte adhesion molecules and extracellular matrix proteins in plastic-embedded human and rat kidneys

    NARCIS (Netherlands)

    van Goor, H; Coers, W; van der Horst, MLC; Suurmeijer, AJH

    2001-01-01

    OBJECTIVE: To study the distribution of cytoskeletal proteins (actin, alpha -actinin, vinculin, beta -tubulin, keratin, vimentin, desmin), adhesion molecules for cell-matrix interations (very later antigens [VLA1-6], beta1, beta2 [CD18], vitronectin receptor [alphav beta3], CD 11b), leukocyte

  19. The influence of matrix properties on growth and morphogenesis of human pancreatic ductal epithelial cells in 3D

    Science.gov (United States)

    Raza, Asad; Ki, Chang Seok; Lin, Chien-Chi

    2013-01-01

    A highly tunable synthetic biomimetic hydrogel platform was developed to study the growth and morphogenesis of pancreatic ductal epithelial cells (PDEC) under the influence of a myriad of instructive cues. A PDEC line, PANC-1, was used as a model system to illustrate the importance of matrix compositions on cell fate determination. PANC-1 is an immortalized ductal epithelial cell line widely used in the study of pancreatic tumor cell behaviors. PANC-1 cells are also increasingly explored as a potential cell source for endocrine differentiation. Thus far, most studies related to PANC-1, among other PDEC lines, are performed on 2D culture surfaces. Here, we evaluated the effect of matrix compositions on PANC-1 cell growth and morphogenesis in 3D. Specifically, PANC-1 cells were encapsulated in PEG-based hydrogels prepared by step-growth thiol-ene photopolymerization. It was found that thiol-ene hydrogels provided a cytocompatible environment for encapsulation and 3D culture of PANC-1 cells. In contrast to a monolayer morphology on 2D culture surfaces, PANC-1 cells formed clusters in 3D thiol-ene hydrogels within 4 days of culture. After culturing for 10 days, however, the growth and structures of these clusters were significantly impacted by gel matrix properties, including sensitivity of the matrix to proteases, stiffness of the matrix, and ECM-mimetic motifs. The use of matrix metalloproteinase (MMP) sensitive linker or the immobilization of fibronectin-derived RGDS ligand in the matrix promoted PANC-1 cell growth and encouraged them to adopt ductal cyst-like structures. On the other hand, the encapsulated cells formed smaller and more compact aggregates in non-MMP responsive gels. The incorporation of laminin-derived YIGSR peptide did not enhance cell growth and caused the cells to form compact aggregates. Immobilized YIGSR also enhanced the expression of epithelial cell markers including β-catenin and E-cadherin. These studies have established PEG

  20. Fibrin glue as agent for sealing corneal and conjunctival wound leaks.

    Science.gov (United States)

    Scalcione, C; Ortiz-Vaquerizas, D; Said, D G; Dua, H S

    2018-02-01

    PurposeTo describe a novel use of fibrin glue in managing leaking blebs and leaking wounds following trauma or surgery.MethodsInterventional case series.ResultsWe report eight patients, including three where intra-operative or immediate post-penetrating keratoplasty recalcitrant leaks from the graft-host junction and/or openings created by the needle pass, were noted. All three had thin recipient beds in the sector of leak. This was managed by intra-cameral injection of fibrin glue in the affected quadrant. This stopped the leak and allowed the defect to heal. One patient of Descemets-stripping-endothelial-keratoplasty had leak from the surgical wound, which was also sealed with fibrin glue. Two patients with leaking glaucoma-surgery-related blebs were treated with intra-bleb injection of fibrin glue to stop the leak. One patient with a penetrating corneal injury with a metal wire had a brisk leak upon removal of the wire. This was sealed with fibrin glue. Another patient of chemical burn with spontaneous leaks was managed by glue injection in the perforations. Transient rise of intraocular pressure in one patient with a leaking bleb was the only adverse event recorded.ConclusionThis novel adaptation of the application of fibrin glue can help to deal with persistent intra-operative, post-operative and traumatic aqueous and air leaks.

  1. Comparative Study of Fibrin Sealent versus Use of Tackers in Inguinal Hernia Repair

    Directory of Open Access Journals (Sweden)

    Wasim Qadir Kar

    2015-09-01

    Full Text Available Purpose: To determine role and benefit of fibrin glue over tackers for mesh fixation in laparoscopic inguinal hernia repair. Backgroud: Mesh fixation by tackers may lead to many complications peroperatively like bleeding, increased hospital stay and overall more cost and later on chronic groin pain. Material and Methods: 60 inguinal hernia with age more than 18 years were taken and were divided in two groups; 30 patient group who underwent TAPP and 30 patient group who underwent TEPP. In 15 patients in both groups tackers were used and in other half fibrin glue was used for fixation of mesh using a 3mm catheter (Duplotip: Baxter Healthcare, which fits the Tisseel syringe. Results: The use of fibrin sealent has a distinct advantage in laparoscopic treatment of inguinal hernias compared with use of tackers as a method of mesh fixation. The use of fibrin sealant reduces the risk of post- and intraoperative complications, such as bleeding, seroma, chronic pain, has a lower incidence of postoperative neuralgia and provides an early faster return to social life. The recurrence rates do not vary much, but the operative time is slightly longer if the preparation time of the fibrin sealant is taken into consideration. In our study, we found a marginal difference in the cost of the two groups, fibrin sealant and stapled tackers. [Cukurova Med J 2015; 40(3.000: 457-465

  2. Human rheumatoid arthritis tissue production of IL-17A drives matrix and cartilage degradation: synergy with tumour necrosis factor-alpha, Oncostatin M and response to biologic therapies.

    LENUS (Irish Health Repository)

    Moran, Ellen M

    2009-01-01

    INTRODUCTION: The aim of this study was to examine IL-17A in patients, following anti-TNF-alpha therapy and the effect of IL-17A on matrix turnover and cartilage degradation. METHODS: IL-17A expression was examined by ELISA and immunohistology in the rheumatoid arthritis (RA) joints. RA whole synovial tissue explant (RA ST), primary synovial fibroblasts (RASFC), human cartilage and chondrocyte cultures were stimulated with IL-17A +\\/- TNF-alpha and Oncostatin M (OSM). Matrix metalloproteinase (MMP) and tissue inhibitor (TIMP-1) were assessed by ELISA and zymography. Cartilage proteoglycan release was assessed histologically by Safranin-O staining. Clinical parameters, IL-17A, MMP\\/TIMP were assessed in patients pre\\/post biologic therapy. RESULTS: IL-17A levels were higher in RA vs osteoarthritis (OA)\\/normal joints (P < 0.05). IL-17A up-regulated MMP-1, -2, -9, and -13 in RA ST, RASFC, cartilage and chondrocyte cultures (P < 0.05). In combination with TNF-alpha and OSM, IL-17A shifted the MMP:TIMP-1 ratio in favor of matrix degradation (all P < 0.05). Cartilage proteoglycan depletion in response to IL-17A was mild; however, in combination with TNF-alpha or OSM showed almost complete proteoglycan depletion. Serum IL-17A was detected in 28% of patients commencing biologic therapy. IL-17A negative patients demonstrated reductions post therapy in serum MMP1\\/TIMP4, MMP3\\/TIMP1 and MMP3\\/TIMP4 ratios and an increase in CS846 (all P < 0.05). No significant changes were observed in IL-17A positive patients. CONCLUSIONS: IL-17A is produced locally in the inflamed RA joint. IL-17A promotes matrix turnover and cartilage destruction, especially in the presence of other cytokines, mimicking the joint environment. IL-17A levels are modulated in vivo, following anti-TNF therapy, and may reflect changes in matrix turnover.

  3. Type VII collagen is enriched in the enamel organic matrix associated with the dentin-enamel junction of mature human teeth.

    Science.gov (United States)

    McGuire, Jacob D; Walker, Mary P; Mousa, Ahmad; Wang, Yong; Gorski, Jeff P

    2014-06-01

    The inner enamel region of erupted teeth is known to exhibit higher fracture toughness and crack growth resistance than bulk phase enamel. However, an explanation for this behavior has been hampered by the lack of compositional information for the residual enamel organic matrix. Since enamel-forming ameloblasts are known to express type VII collagen and type VII collagen null mice display abnormal amelogenesis, the aim of this study was to determine whether type VII collagen is a component of the enamel organic matrix at the dentin-enamel junction (DEJ) of mature human teeth. Immunofluorescent confocal microscopy of demineralized tooth sections localized type VII collagen to the organic matrix surrounding individual enamel rods near the DEJ. Morphologically, immunoreactive type VII collagen helical-bundles resembled the gnarled-pattern of enamel rods detected by Coomassie Blue staining. Western blotting of whole crown or enamel matrix extracts also identified characteristic Mr=280 and 230 kDa type VII dimeric forms, which resolved into 75 and 25 kDa bands upon reduction. As expected, the collagenous domain of type VII collagen was resistant to pepsin digestion, but was susceptible to purified bacterial collagenase. These results demonstrate the inner enamel organic matrix in mature teeth contains macromolecular type VII collagen. Based on its physical association with the DEJ and its well-appreciated capacity to complex with other collagens, we hypothesize that enamel embedded type VII collagen fibrils may contribute not only to the structural resilience of enamel, but may also play a role in bonding enamel to dentin. Copyright © 2014 Elsevier Inc. All rights reserved.

  4. Development of a tissue-engineered human oral mucosa equivalent based on an acellular allogeneic dermal matrix: a preliminary report of clinical application to burn wounds.

    Science.gov (United States)

    Iida, Takuya; Takami, Yoshihiro; Yamaguchi, Ryo; Shimazaki, Shuji; Harii, Kiyonori

    2005-01-01

    Tissue-engineered skin equivalents composed of epidermal and dermal components have been widely investigated for coverage of full-thickness skin defects. We developed a tissue-engineered oral mucosa equivalent based on an acellular allogeneic dermal matrix and investigated its characteristics. We also tried and assessed its preliminary clinical application. Human oral mucosal keratinocytes were separated from a piece of oral mucosa and cultured in a chemically-defined medium. The keratinocytes were seeded on to the acellular allogeneic dermal matrix and cultured. Histologically, the mucosa equivalent had a well-stratified epithelial layer. Immunohistochemical study showed that it was similar to normal oral mucosa. We applied this equivalent in one case with an extensive burn wound. The equivalent was transplanted three weeks after the harvest of the patient's oral mucosa and about 30% of the graft finally survived. We conclude that this new oral mucosa equivalent could become a therapeutic option for the treatment of extensive burns.

  5. Matrix production and organization by endothelial colony forming cells in mechanically strained engineered tissue constructs.

    Directory of Open Access Journals (Sweden)

    Nicky de Jonge

    Full Text Available AIMS: Tissue engineering is an innovative method to restore cardiovascular tissue function by implanting either an in vitro cultured tissue or a degradable, mechanically functional scaffold that gradually transforms into a living neo-tissue by recruiting tissue forming cells at the site of implantation. Circulating endothelial colony forming cells (ECFCs are capable of differentiating into endothelial cells as well as a mesenchymal ECM-producing phenotype, undergoing Endothelial-to-Mesenchymal-transition (EndoMT. We investigated the potential of ECFCs to produce and organize ECM under the influence of static and cyclic mechanical strain, as well as stimulation with transforming growth factor β1 (TGFβ1. METHODS AND RESULTS: A fibrin-based 3D tissue model was used to simulate neo-tissue formation. Extracellular matrix organization was monitored using confocal laser-scanning microscopy. ECFCs produced collagen and also elastin, but did not form an organized matrix, except when cultured with TGFβ1 under static strain. Here, collagen was aligned more parallel to the strain direction, similar to Human Vena Saphena Cell-seeded controls. Priming ECFC with TGFβ1 before exposing them to strain led to more homogenous matrix production. CONCLUSIONS: Biochemical and mechanical cues can induce extracellular matrix formation by ECFCs in tissue models that mimic early tissue formation. Our findings suggest that priming with bioactives may be required to optimize neo-tissue development with ECFCs and has important consequences for the timing of stimuli applied to scaffold designs for both in vitro and in situ cardiovascular tissue engineering. The results obtained with ECFCs differ from those obtained with other cell sources, such as vena saphena-derived myofibroblasts, underlining the need for experimental models like ours to test novel cell sources for cardiovascular tissue engineering.

  6. Osthole Suppresses the Migratory Ability of Human Glioblastoma Multiforme Cells via Inhibition of Focal Adhesion Kinase-Mediated Matrix Metalloproteinase-13 Expression

    Directory of Open Access Journals (Sweden)

    Cheng-Fang Tsai

    2014-03-01

    Full Text Available Glioblastoma multiforme (GBM is the most common type of primary and malignant tumor occurring in the adult central nervous system. GBM often invades surrounding regions of the brain during its early stages, making successful treatment difficult. Osthole, an active constituent isolated from the dried C. monnieri fruit, has been shown to suppress tumor migration and invasion. However, the effects of osthole in human GBM are largely unknown. Focal adhesion kinase (FAK is important for the metastasis of cancer cells. Results from this study show that osthole can not only induce cell death but also inhibit phosphorylation of FAK in human GBM cells. Results from this study show that incubating GBM cells with osthole reduces matrix metalloproteinase (MMP-13 expression and cell motility, as assessed by cell transwell and wound healing assays. This study also provides evidence supporting the potential of osthole in reducing FAK activation, MMP-13 expression, and cell motility in human GBM cells.

  7. Matrix Metalloproteinase Enzyme Family

    Directory of Open Access Journals (Sweden)

    Ozlem Goruroglu Ozturk

    2013-04-01

    Full Text Available Matrix metalloproteinases play an important role in many biological processes such as embriogenesis, tissue remodeling, wound healing, and angiogenesis, and in some pathological conditions such as atherosclerosis, arthritis and cancer. Currently, 24 genes have been identified in humans that encode different groups of matrix metalloproteinase enzymes. This review discuss the members of the matrix metalloproteinase family and their substrate specificity, structure, function and the regulation of their enzyme activity by tissue inhibitors. [Archives Medical Review Journal 2013; 22(2.000: 209-220

  8. Rodent neonatal germinal matrix hemorrhage mimics the human brain injury, neurological consequences, and post-hemorrhagic hydrocephalus

    OpenAIRE

    Lekic, Tim; Manaenko, Anatol; Rolland, William; Krafft, Paul R.; Peters, Regina; Hartman, Richard E.; Altay, Orhan; Tang, Jiping; Zhang, John H.

    2012-01-01

    Germinal matrix hemorrhage (GMH) is the most common neurological disease of premature newborns. GMH causes neurological sequelae such as cerebral palsy, post-hemorrhagic hydrocephalus, and mental retardation. Despite this, there is no standardized animal model of spontaneous GMH using newborn rats to depict the condition. We asked whether stereotactic injection of collagenase type VII (0.3 U) into the ganglionic eminence of neonatal rats would reproduce the acute brain injury, gliosis, hydroc...

  9. Treatment of venous ulcers with fibrin sealant derived from snake venom

    Directory of Open Access Journals (Sweden)

    MAN Gatti

    2011-01-01

    Full Text Available Venous ulcers of the lower limbs complicated by infection or chronicity represent a serious public health problem. The elevated number of those afflicted burdens the health services, interferes in quality of life and causes absenteeism. Although there are 2,500 items on the market, ranging from the simplest dressing up to the most complex types of dressing, treatment remains a challenge. Among the substances used, fibrin sealant is the one that promotes diminution of bacterial colonization and of edema, controls hemorrhaging, alters the pain threshold by protecting the nerve endings, hydrates the wound bed and forms granulation tissue that favors healing. Its disadvantages include higher cost and utilization of human fibrinogen that can transmit infectious diseases. The Center for the Study of Venoms and Venomous Animals (CEVAP at São Paulo State University (UNESP developed a new sealant made up of fibrinogen extracted from large animals and from an enzyme obtained from snake venom. The present study, developed in the Health Education Clinic (CEPS of Sacred Heart University (USC aimed to evaluate the effect of the new sealant on the healing process of venous ulcers in 24 adult patients, seven of whom were male and 17 female. Two study groups were formed as follows: Group 1 (G1 - control group of 11 patients treated with essential fatty acid (EFA and Unna's boot, and Group 2 (G2 - 13 patients treated with essential fatty acid (EFA, fibrin sealant and Unna's boot. The follow-up lasted eight weeks and the sealant was applied at only the first and fourth weeks. The results showed that Group 2 presented worse lesion conditions as to healing, but, when comparing the two groups, it was noteworthy that the the sealant was effective in healing venous ulcers. There is evidence that the new sealant is recommended for leg ulcers with the following advantages: ease of application, preparation of the wound bed, diminution of pain and a higher number of

  10. Polyethylene glycol–polylactic acid nanoparticles modified with cysteine–arginine–glutamic acid–lysine–alanine fibrin-homing peptide for glioblastoma therapy by enhanced retention effect

    Directory of Open Access Journals (Sweden)

    Wu J

    2014-11-01

    Full Text Available Junzhu Wu,1,2,* Jingjing Zhao,1,3,* Bo Zhang,1 Yong Qian,1 Huile Gao,1 Yuan Yu,1 Yan Wei,1 Zhi Yang,1 Xinguo Jiang,1 Zhiqing Pang1 1Key Laboratory of Smart Drug Delivery, Ministry of Education, Department of Pharmaceutics, School of Pharmacy, Fudan University, Shanghai, 2School of Pharmacy, Dali University, Xiaguan, 3School of Pharmacy, China Pharmaceutical University, Nanjing, People’s Republic of China *These authors contributed equally to this work Abstract: For a nanoparticulate drug-delivery system, crucial challenges in brain-glioblastoma therapy are its poor penetration and retention in the glioblastoma parenchyma. As a prevailing component in the extracellular matrix of many solid tumors, fibrin plays a critical role in the maintenance of glioblastoma morphology and glioblastoma cell differentiation and proliferation. We developed a new drug-delivery system by conjugating polyethylene glycol–polylactic acid nanoparticles (NPs with cysteine–arginine–glutamic acid–lysine–alanine (CREKA; TNPs, a peptide with special affinity for fibrin, to mediate glioblastoma-homing and prolong NP retention at the tumor site. In vitro binding tests indicated that CREKA significantly enhanced specific binding of NPs with fibrin. In vivo fluorescence imaging of glioblastoma-bearing nude mice, ex vivo brain imaging, and glioblastoma distribution demonstrated that TNPs had higher accumulation and longer retention in the glioblastoma site over unmodified NPs. Furthermore, pharmacodynamic results showed that paclitaxel-loaded TNPs significantly prolonged the median survival time of intracranial U87 glioblastoma-bearing nude mice compared with controls, Taxol, and NPs. These findings suggested that TNPs were able to target the glioblastoma and enhance retention, which is a valuable strategy for tumor therapy. Keywords: CREKA peptide, nanoparticles, retention effect, paclitaxel, glioblastoma

  11. The impact of the centrifuge characteristics and centrifugation protocols on the cells, growth factors, and fibrin architecture of a leukocyte- and platelet-rich fibrin (L-PRF) clot and membrane.

    Science.gov (United States)

    Dohan Ehrenfest, David M; Pinto, Nelson R; Pereda, Andrea; Jiménez, Paula; Corso, Marco Del; Kang, Byung-Soo; Nally, Mauricio; Lanata, Nicole; Wang, Hom-Lay; Quirynen, Marc

    2018-03-01

    -PRF. All centrifugations were done using the original L-PRF centrifuge (Intra-Spin), as recommended by the two manufacturers. Half of the membranes were placed individually in culture media and transferred in a new tube at seven experimental times (up to 7 days). The releases of transforming growth factor β-1 (TGFβ-1), platelet derived growth factor AB (PDGF-AB), vascular endothelial growth factor (VEGF) and bone morphogenetic protein 2 (BMP-2) were quantified using ELISA kits at these seven experimental times. The remaining membranes were used to evaluate the initial quantity of growth factors of the L-PRF and A-PRF membranes, through forcible extraction. Very significant differences in the level of vibrations at each rotational speed were observed between the four tested centrifuges. The original L-PRF centrifuge (Intra-Spin) was by far the most stable machine in all configurations and always remained under the threshold of resonance, unlike the three other tested machines. At the classical speed of production of L-PRF, the level of undesirable vibrations on the original centrifuge was between 4.5 and 6 times lower than with other centrifuges. Intra-Spin showed the lowest temperature of the tubes. A-PRF and Salvin were both associated with a significant increase in temperature in the tube. Intra-Spin produced the heaviest clot and quantity of exudate among the four techniques. A-PRF and LW produced much lighter, shorter and narrower clots and membranes than the two other centrifuges. Light microscopy analysis showed relatively similar features for all L-PRF types (concentration of cell bodies in the first half). However, SEM illustrated considerable differences between samples. The original Intra-Spin L-PRF showed a strongly polymerized thick fibrin matrix and all cells appeared alive with a normal shape, including the textured surface aspect of activated lymphocytes. The A-PRF, Salvin and LW PRF-like membranes presented a lightly polymerized slim fibrin gel and most

  12. Detection and quantification of neurotensin in human brain tissue by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    DEFF Research Database (Denmark)

    Gobom, J; Kraeuter, K O; Persson, R

    2000-01-01

    A method was developed for mass spectrometric detection of neurotensin (NT)-like immunoreactivity and quantification of NT in human brain tissue. The method is based on immunoprecipitation followed by analysis using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF......-MS). The identity of the major component of the immunoprecipitates as neurotensin was confirmed by fragment ion analysis on an electrospray ionization quadrupole time-of-flight instrument. MALDI-TOF-MS quantification of NT was achieved using stable-isotope-labeled NT as the internal standard, yielding an error...

  13. Safety and hemostatic efficacy of fibrin pad in partial nephrectomy: Results of an open-label Phase I and a randomized, standard-of-care-controlled Phase I/II study

    Directory of Open Access Journals (Sweden)

    Nativ Ofer

    2012-11-01

    Full Text Available Abstract Background Bleeding severity, anatomic location, tissue characteristics, and visibility are common challenges encountered while managing intraoperative bleeding, and conventional hemostatic measures (suture, ligature, and cautery may sometimes be ineffective or impractical. While topical absorbable hemostats (TAH are useful hemostatic adjuvants, each TAH has associated disadvantages. Methods We evaluated the safety and hemostatic efficacy of a new advanced biologic combination product―fibrin pad―to potentially address some gaps associated with TAHs. Fibrin pad was assessed as adjunctive hemostat in open partial nephrectomy in single-center, open-label, Phase I study (N = 10, and as primary hemostat in multicenter, single-blind, randomized, standard-of-care (SOC-controlled Phase I/II study (N = 7 in Israel. It was used to control mild-to-moderate bleeding in Phase I and also spurting arterial bleeding in Phase I/II study. Phase I study assessed safety and Phase I/II study, proportion of successes at 10 min following randomization, analyzed by Fisher exact tests at 5% significance level. Results Phase I (N = 10: All patients completed the study. Hemostasis was achieved within 3–4 min (average = 3.1 min of a single application in all patients. Fibrin pad was found to be safe for human use, with no product-related adverse events reported. Phase I/II (N = 7: Hemostatic success at 10 min (primary endpoint was achieved in 3/4 patients treated with fibrin pad versus 0/3 patients treated with SOC. No clinically significant change in laboratory or coagulation parameters was recorded, except a case of post-procedural hemorrhage with fibrin pad, which was considered serious and related to the fibrin pad treatment, and required re-operation. Although Data Safety Monitoring Board authorized trial continuation, the sponsor decided against proceeding toward an indication for primary treatment of severe arterial

  14. Co-effects of matrix low elasticity and aligned topography on stem cell neurogenic differentiation and rapid neurite outgrowth

    Science.gov (United States)

    Yao, Shenglian; Liu, Xi; Yu, Shukui; Wang, Xiumei; Zhang, Shuming; Wu, Qiong; Sun, Xiaodan; Mao, Haiquan

    2016-05-01

    The development of novel biomaterials that deliver precise regulatory signals to direct stem cell fate for nerve regeneration is the focus of current intensive research efforts. In this study, a hierarchically aligned fibrillar fibrin hydrogel (AFG) that was fabricated through electrospinning and the concurrent molecular self-assembly process mimics both the soft and oriented features of nerve tissue, thus providing hybrid biophysical cues to instruct cell behavior in vitro and in vivo. The electrospun hydrogels were examined by scanning electron microscopy (SEM), polarized light microscopy, small angle X-ray scattering assay and atomic force microscopy (AFM), showing a hierarchically linear-ordered structure from the nanoscale to the macroscale with a soft elastic character (elasticity ~1 kPa). We found that this low elasticity and aligned topography of AFG exhibit co-effects on promoting the neurogenic differentiation of human umbilical cord mesenchymal stem cells (hUMSCs) in comparison to random fibrin hydrogel (RFG) and tissue culture plate (TCP) control after two week cell culture in growth medium lacking supplementation with soluble neurogenic induction factors. In addition, AFG also induces dorsal root ganglion (DRG) neurons to rapidly project numerous long neurite outgrowths longitudinally along the AFG fibers for a total neurite extension distance of 1.96 mm in three days in the absence of neurotrophic factor supplementation. Moreover, the AFG implanted in a rat T9 dorsal hemisection spinal cord injury model was found to promote endogenous neural cell fast migration and axonal invasion along AFG fibers, resulting in aligned tissue cables in vivo. Our results suggest that matrix stiffness and aligned topography may instruct stem cell neurogenic differentiation and rapid neurite outgrowth, providing great promise for biomaterial design for applications in nerve regeneration.The development of novel biomaterials that deliver precise regulatory signals to

  15. Identification of p63+ keratinocyte progenitor cells in circulation and their matrix-directed differentiation to epithelial cells.

    Science.gov (United States)

    Nair, Renjith P; Krishnan, Lissy K

    2013-04-11

    In the event of chronic diabetes or burn wounds, accomplishing skin regeneration is a major concern. Autologous skin grafting is the most effective remedy, but the tissue harvest may create more nonhealing wounds. Currently available skin substitutes have a limited clinical outcome because of immune reactions arising from the xenobiotic scaffold or allogenous cells. Autologous stem cells that can be collected without an additional injury may be a viable option for skin-tissue engineering. Presence of a low number of keratinocyte progenitor cells (KPCs) within the peripheral blood mononuclear cell (PBMNC) population has been indicated. Identification, isolation, expansion, and differentiation of KPCs is necessary before they are considered for skin regeneration, which is the focus of this study. Culture of isolated human PBMNCs on a cell-specific matrix was carried out to induce differentiation of KPCs. Flow cytometry and reverse transcriptase polymerase chain reaction were done for epithelial stem cell marker p63 and lineage markers cytokeratin 5 and cytokeratin 14, to track differentiation. Proliferation was confirmed by quantifying the proliferating cell nuclear antigen-expressing cells. Immunostaining with epithelial cell markers, involucrin and filaggrin, was carried out to establish terminal differentiation. Microscopic analysis confirmed growth and survival of KPCs on the dermal fibroblast monolayer and on a transplantable fibrin sheet. We demonstrated that KPCs are p63(+) and CD34-. The specifically designed composition of the extracellular matrix was found to support selective adhesion, proliferation, and differentiation of p63(+) KPCs. The PBMNC culture for 12 days under controlled conditions resulted in a homogenous population that expressed cytokeratins, and >90% of the cells were found to proliferate. Subculture for 5 days resulted in expression of filaggrin and involucrin, suggesting terminal differentiation. Transfer of matrix-selected KPCs to a

  16. Evaluation of dermal-epidermal skin equivalents ('composite-skin') of human keratinocytes in a collagen-glycosaminoglycan matrix(Integra artificial skin).

    Science.gov (United States)

    Kremer, M; Lang, E; Berger, A C

    2000-09-01

    Integra artificial skin (Integra LifeSciences Corp., Plainsboro, NJ, USA) is a dermal template consisting of bovine collagen, chondroitin-6-sulphate and a silastic membrane manufactured as Integra. This product has gained widespread use in the clinical treatment of third degree burn wounds and full thickness skin defects of different aetiologies. The product was designed to significantly reduce the time needed to achieve final wound closure in the treatment of major burn wounds, to optimise the sparse autologous donor skin resources and to improve the durable mechanical quality of the skin substitute. The clinical procedure requires two stages. The first step creates a self neodermis, the second creates a self epidermis on the neodermis. However, it is desirable to cover major burn wounds early in a single step by a skin substitute consisting of a dermal equivalent seeded in vitro with autologous keratinocytes ('composite-skin') out of which a full thickness skin develops in vivo.The goal of this experimental study was to develop a method to integrate human keratinocytes in homogeneous distribution and depth into Integra Artificial Skin. The seeded cell-matrix composites were grafted onto athymic mice in order to evaluate their potential to reconstitute a human epidermis in vivo. We were able to demonstrate that the inoculated human keratinocytes reproducibly displayed a homogeneous pattern of distribution, adherence, proliferation and confluence. The cell-matrix composites grafted in this model exhibited good wound adherence, complete healing, minor wound contraction and had the potential to reconstitute an elastic, functional and durable human skin. Histologically we were able to show that the inoculated human keratinocytes in vivo colonised the matrix in a histomorphologically characteristic epidermal pattern (keratomorula, keratinocyte bubbling) and developed a persisting, stratified, keratinising epidermis which immunohistologically proved to be of human

  17. Scaffold composition affects cytoskeleton organization, cell-matrix interaction and the cellular fate of human mesenchymal stem cells upon chondrogenic differentiation.

    Science.gov (United States)

    Li, Yuk Yin; Choy, Tze Hang; Ho, Fu Chak; Chan, Pui Barbara

    2015-06-01

    The stem cell niche, or microenvironment, consists of soluble, matrix, cell and mechanical factors that together determine the cellular fates and/or differentiation patterns of stem cells. Collagen and glycosaminoglycans (GAGs) are important scaffolding materials that can mimic the natural matrix niche. Here, we hypothesize that imposing changes in the scaffold composition or, more specifically, incorporating GAGs into the collagen meshwork, will affect the morphology, cytoskeletal organization and integrin expression profiles, and hence the fate of human mesenchymal stem cells (MSCs) upon the induction of differentiation. Using chondrogenesis as an example, we microencapsulated MSCs in three scaffold systems that had varying matrix compositions: collagen alone (C), aminated collagen (AC) and aminated collagen with GAGs (ACG). We then induced the MSCs to differentiate toward a chondrogenic lineage, after which, we characterized the cell viability and morphology, as well as the level of cytoskeletal organization and the integrin expression profile. We also studied the fate of the MSCs by evaluating the major chondrogenic markers at both the gene and protein level. In C, MSC chondrogenesis was successfully induced and MSCs that spread in the scaffolds had a clear actin cytoskeleton; they expressed integrin α2β1, α5 and αv; promoted sox9 nuclear localization transcription activation; and upregulated the expression of chondrogenic matrix markers. In AC, MSC chondrogenesis was completely inhibited but the scaffold still supported cell survival. The MSCs did not spread and they had no actin cytoskeleton; did not express integrin α2 or αv; they failed to differentiate into chondrogenic lineage cells even on chemical induction; and there was little colocalization or functional interaction between integrin α5 and fibronectin. In ACG, although the MSCs did not express integrin α2, they did express integrin αv and there was strong co-localization and hence functional

  18. Efficacy of leukocyte- and platelet-rich fibrin in wound healing: a randomized controlled clinical trial.

    Science.gov (United States)

    Chignon-Sicard, Bérengère; Georgiou, Charalambos A; Fontas, Eric; David, Sylvain; Dumas, Pierre; Ihrai, Tarik; Lebreton, Elisabeth

    2012-12-01

    Application of platelet concentrates to wounds could speed healing. Leukocyte- and platelet-rich fibrin, a relatively recent development, stands out from the other preparations. This prospective, randomized, controlled clinical trial studied the rate of healing of postoperative hand wounds after a single application of leukocyte- and platelet-rich fibrin. Eligible patients were healthy individuals older than 18 years who had been scheduled for elective McCash (open palm) surgery for Dupuytren disease at the Plastic and Hand Surgery Department of Nice's University Hospital between August of 2007 and February of 2010. The control group received the reference care of petroleum jelly mesh (Vaselitulle), and test patients had leukocyte- and platelet-rich fibrin applied. The primary endpoint was healing delay measured in postoperative days. Secondary endpoints included pain, bleeding, and wound exudate. The trial was carried out as a single-blind trial. Among the 68 randomized patients, 33 patients in the leukocyte- and platelet-rich fibrin group and 31 in the Vaselitulle group were analyzed. Primary endpoint analysis showed a median healing delay of 24 days (interquartile range, 18 to 28 days) for the fibrin group and 29 days (interquartile range, 26 to 35 days) for the Vaselitulle group (p = 0.014, log-rank test). Postoperative pain assessment, bleeding, and exudate were always lower for the fibrin group, but not significantly so. The authors trial demonstrates that a single leukocyte- and platelet-rich fibrin application on fresh postoperative hand wounds shows a median improvement of 5 days in comparison with the standard treatment. Therapeutic, II.

  19. Tracheal anastomosis with the diode laser and fibrin tissue adhesive: an in vitro and in vivo investigation.

    Science.gov (United States)

    Gleich, L L; Wang, Z; Pankratov, M M; Aretz, H T; Shapshay, S M

    1995-05-01

    Absorbable sutures have been advocated for tracheal anastomosis to reduce fibrosis and foreign body reaction leading to recurrent stenosis. Fibrin tissue adhesive (FTA) and diode laser welding with indocyanine green-dyed fibrinogen were evaluated in tracheal anastomosis to reduce the number of sutures and to improve healing. In vitro studies demonstrated strong anastomoses with a combination of laser welding and FTA with minimal tissue damage. In a controlled in vivo study, circumferential resections of canine tracheas were repaired with laser welding and FTA augmented with a few stay sutures. These anastomoses had less fibrosis and tissue damage than anastomoses in control animals repaired with sutures alone. This study supports investigation of laser welding and FTA in human beings for tracheal anastomosis and other procedures in which suturing may be difficult.

  20. Fibroblast-Derived Extracellular Matrix Induces Chondrogenic Differentiation in Human Adipose-Derived Mesenchymal Stromal/Stem Cells in Vitro

    Directory of Open Access Journals (Sweden)

    Kevin Dzobo

    2016-08-01

    Full Text Available 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.

  1. Matrix calculus

    CERN Document Server

    Bodewig, E

    1959-01-01

    Matrix Calculus, Second Revised and Enlarged Edition focuses on systematic calculation with the building blocks of a matrix and rows and columns, shunning the use of individual elements. The publication first offers information on vectors, matrices, further applications, measures of the magnitude of a matrix, and forms. The text then examines eigenvalues and exact solutions, including the characteristic equation, eigenrows, extremum properties of the eigenvalues, bounds for the eigenvalues, elementary divisors, and bounds for the determinant. The text ponders on approximate solutions, as well

  2. Pancreatic fistula after laparoscopic splenectomy in patients with hypersplenism due to liver cirrhosis: effect of fibrin glue and polyglycolic acid felt on prophylaxis of postoperative complications.

    Science.gov (United States)

    Tsutsumi, Norifumi; Tomikawa, Morimasa; Akahoshi, Tomohiko; Kawanaka, Hirofumi; Ota, Mitsuhiko; Sakaguchi, Yoshihisa; Kusumoto, Tetsuya; Ikejiri, Koji; Hashizume, Makoto; Maehara, Yoshihiko

    2016-11-01

    This study aimed to determine the effect of fibrin glue and polyglycolic acid (PGA) felt on prevention of pancreatic fistula (PF) after laparoscopic splenectomy in patients with hypersplenism due to liver cirrhosis. Fifty consecutive patients were enrolled in this prospective study. Twenty-three patients underwent laparoscopic splenectomy with a fibrin sheet (fibrin sheet group). The sealing ability of each treatment was evaluated by an ex vivo pressure test model. Based on the results from ex vivo experiments, 27 patients received prophylaxis using fibrin glue and PGA felt (PGA with fibrin group). The primary endpoint was the incidence of PF. Significantly more (5, 22%) patients developed PF in the fibrin sheet group than in the PGA with fibrin group (0%, P = .037). Our new application of fibrin glue and PGA felt is an effective prophylactic procedure for preventing development of PF after laparoscopic splenectomy. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Co-effects of matrix low elasticity and aligned topography on stem cell neurogenic differentiation and rapid neurite outgrowth.

    Science.gov (United States)

    Yao, Shenglian; Liu, Xi; Yu, Shukui; Wang, Xiumei; Zhang, Shuming; Wu, Qiong; Sun, Xiaodan; Mao, Haiquan

    2016-05-21

    The development of novel biomaterials that deliver precise regulatory signals to direct stem cell fate for nerve regeneration is the focus of current intensive research efforts. In this study, a hierarchically aligned fibrillar fibrin hydrogel (AFG) that was fabricated through electrospinning and the concurrent molecular self-assembly process mimics both the soft and oriented features of nerve tissue, thus providing hybrid biophysical cues to instruct cell behavior in vitro and in vivo. The electrospun hydrogels were examined by scanning electron microscopy (SEM), polarized light microscopy, small angle X-ray scattering assay and atomic force microscopy (AFM), showing a hierarchically linear-ordered structure from the nanoscale to the macroscale with a soft elastic character (elasticity ∼1 kPa). We found that this low elasticity and aligned topography of AFG exhibit co-effects on promoting the neurogenic differentiation of human umbilical cord mesenchymal stem cells (hUMSCs) in comparison to random fibrin hydrogel (RFG) and tissue culture plate (TCP) control after two week cell culture in growth medium lacking supplementation with soluble neurogenic induction factors. In addition, AFG also induces dorsal root ganglion (DRG) neurons to rapidly project numerous long neurite outgrowths longitudinally along the AFG fibers for a total neurite extension distance of 1.96 mm in three days in the absence of neurotrophic factor supplementation. Moreover, the AFG implanted in a rat T9 dorsal hemisection spinal cord injury model was found to promote endogenous neural cell fast migration and axonal invasion along AFG fibers, resulting in aligned tissue cables in vivo. Our results suggest that matrix stiffness and aligned topography may instruct stem cell neurogenic differentiation and rapid neurite outgrowth, providing great promise for biomaterial design for applications in nerve regeneration.

  4. A human pericardium biopolymeric scaffold for autologous heart valve tissue engineering: cellular and extracellular matrix structure and biomechanical properties in comparison with a normal aortic heart valve.

    Science.gov (United States)

    Straka, Frantisek; Schornik, David; Masin, Jaroslav; Filova, Elena; Mirejovsky, Tomas; Burdikova, Zuzana; Svindrych, Zdenek; Chlup, Hynek; Horny, Lukas; Daniel, Matej; Machac, Jiri; Skibová, Jelena; Pirk, Jan; Bacakova, Lucie

    2018-04-01

    The objective of our study was to compare the cellular and extracellular matrix (ECM) structure and the biomechanical properties of human pericardium (HP) with the normal human aortic heart valve (NAV). HP tissues (from 12 patients) and NAV samples (from 5 patients) were harvested during heart surgery. The main cells in HP were pericardial interstitial cells, which are fibroblast-like cells of mesenchymal origin similar to the valvular interstitial cells in NAV tissue. The ECM of HP had a statistically significantly (p structures of the two tissues, the dense part of fibrous HP (49 ± 2%) and the lamina fibrosa of NAV (47 ± 4%), was similar. In both tissues, the secant elastic modulus (Es) was significantly lower in the transversal direction (p structure and has the biomechanical properties required for a tissue from which an autologous heart valve replacement may be constructed.

  5. Surgical Outcomes of Deep Superior Sulcus Augmentation Using Acellular Human Dermal Matrix in Anophthalmic or Phthisis Socket.

    Science.gov (United States)

    Cho, Won-Kyung; Jung, Su-Kyung; Paik, Ji-Sun; Yang, Suk-Woo

    2016-07-01

    Patients with anophthalmic or phthisis socket suffer from cosmetic problems. To resolve those problems, the authors present the surgical outcomes of deep superior sulcus (DSS) augmentation using acellular dermal matrix in patients with anophthalmic or phthisis socket. The authors retrospectively reviewed anophthalmic or phthisis patients who underwent surgery for DSS augmentation using acellular dermal matrix. To evaluate surgical outcomes, the authors focused on 3 aspects: the possibility of wearing contact prosthesis, the degree of correction of the DSS, and any surgical complications. The degree of correction of DSS was classified as excellent: restoration of superior sulcus enough to remove sunken sulcus shadow; fair: gain of correction effect but sunken shadow remained; or fail: no effect of correction at all. Ten eyes of 10 patients were included. There was a mean 21.3 ± 37.1-month period from evisceration or enucleation to the operation for DSS augmentation. All patients could wear contact prosthesis after the operation (100%). The degree of correction was excellent in 8 patients (80%) and fair in 2. Three of 10 (30%) showed complications: eyelid entropion, upper eyelid multiple creases, and spontaneous wound dehiscence followed by inflammation after stitch removal. Uneven skin surface and paresthesia in the forehead area of the affected eye may be observed after surgery. The overall surgical outcomes were favorable, showing an excellent degree of correction of DSS and low surgical complication rates. This procedure is effective for patients who have DSS in the absence or atrophy of the eyeball.

  6. Intracellular Wnt/Beta-Catenin Signaling Underlying 17beta-Estradiol-Induced Matrix Metalloproteinase 9 Expression in Human Endometriosis.

    Science.gov (United States)

    Zhang, Ling; Xiong, Wenqian; Xiong, Yao; Liu, Hengwei; Li, Na; Du, Yu; Liu, Yi

    2016-03-01

    Extracellular matrix remodeling is necessary for ectopic endometrium implantation. Many studies have shown an increased expression of matrix metalloproteinase 9 (MMP9) in the ectopic endometrium of endometriosis. However, the signaling pathways and cellular effects related to this process remain incompletely elucidated. The objective of our study was to investigate the association between MMP9 and the Wnt signaling pathway under the regulation of 17beta-estradiol (E2) in endometrial stromal cells. We found that MMP9 was elevated in tissues from women with endometriosis compared with normal women. Furthermore, MMP9 and beta-catenin increased concurrently in a time- and dose-dependent manner after E2 treatment. To clarify the relationship between MMP9 and beta-catenin, we performed luciferase promoter reporter and chromatin immunoprecipitation assays. A beta-catenin/TCF3/LEF1 complex bound to a specific site on the MMP9 promoter that promoted MMP9 gene and protein expression. The promotion of MMP9 by the Wnt signaling pathway under the regulation of E2 may contribute to the pathophysiology of this disease. © 2016 by the Society for the Study of Reproduction, Inc.

  7. Ultrastructure of collagen fibers and distribution of extracellular matrix in the temporomandibular disk of the human fetus and adult.

    Science.gov (United States)

    Takahashi, H; Sato, I

    2001-12-01

    We quantitatively examined the distribution of these differences in extracellular matrices (collagen types I, III, and fibronectin) and elastic fibers under confocal laser scanning microscopy and electron scanning microscopy in terms of their contribution to the mechanics of the TMJ during development and in adults. Elastic fibers were found in the anterior and posterior bands in adults aged 40 years, and a few elastic fibers in the anterior band of the disk in adults aged 80 to 90 years. The extracellular matrix contents of the TMJ disk are shown in various detected levels in the anterior, intermediate, posterior bands of TMJ disk. During development, collagen fibers are arranged in a complex fashion from 28 weeks' gestation. These ultrastructures of the embryonic TMJ are resembled to that of adults aged the 40s, however the difference in extracellular matrix distribution found in embryonic stages and adults. They might reflect the differences in function between mastication and sucking or the changes in shape and form as results of functional disorders of the TMJ.

  8. The potential of 3-dimensional construct engineered from poly(lactic-co-glycolic acid)/fibrin hybrid scaffold seeded with bone marrow mesenchymal stem cells for in vitro cartilage tissue engineering.

    Science.gov (United States)

    Abdul Rahman, Rozlin; Mohamad Sukri, Norhamiza; Md Nazir, Noorhidayah; Ahmad Radzi, Muhammad Aa'zamuddin; Zulkifly, Ahmad Hafiz; Che Ahmad, Aminudin; Hashi, Abdurezak Abdulahi; Abdul Rahman, Suzanah; Sha'ban, Munirah

    2015-08-01

    Articular cartilage is well known for its simple uniqueness of avascular and aneural structure that has limited capacity to heal itself when injured. The use of three dimensional construct in tissue engineering holds great potential in regenerating cartilage defects. This study evaluated the in vitro cartilaginous tissue formation using rabbit's bone marrow mesenchymal stem cells (BMSCs)-seeded onto poly(lactic-co-glycolic acid) PLGA/fibrin and PLGA scaffolds. The in vitro cartilaginous engineered constructs were evaluated by gross inspection, histology, cell proliferation, gene expression and sulphated glycosaminoglycan (sGAG) production at week 1, 2 and 3. After 3 weeks of culture, the PLGA/fibrin construct demonstrated gross features similar to the native tissue with smooth, firm and glistening appearance, superior histoarchitectural and better cartilaginous extracellular matrix compound in concert with the positive glycosaminoglycan accumulation on Alcian blue. Significantly higher cell proliferation in PLGA/fibrin construct was noted at day-7, day-14 and day-21 (ptissue engineered cartilage. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Stability of Beriplast P fibrin sealant: storage and reconstitution.

    Science.gov (United States)

    Eberhard, Ulrich; Broder, Martin; Witzke, Günther

    2006-04-26

    This study was performed to investigate the stability of Beriplast P fibrin sealant (FS) across a range of storage conditions, both pre- and post-reconstitution. Storage stability of the FS was evaluated during long-term refrigeration (24 months) with or without interim storage at elevated temperatures (40 degrees C for 1 week and 25 degrees C for 1 and 3 months). Stability of individual FS components was assessed by measuring: fibrinogen content, Factor XIII activity (FXIII), thrombin activity and aprotinin potency. The package integrity of each component was also checked (sterility testing, moisture content and pH). Storage stability was also evaluated by testing the reconstituted product for adhesion (tearing force testing after mixing the solutions) and sterility. Reconstitution stability was evaluated following 3-months' storage, for up to 50 h post-reconstitution using the same tests as for the storage stability investigations. Pre-defined specifications were met for fibrinogen content, Factor XIII activity, and thrombin activity, demonstrating storage stability. Package integrity and the functionality and sterility of the reconstituted product were confirmed throughout. Reconstitution stability was demonstrated for up to 50 h following reconstitution, in terms of both tearing force and sterility tests. In conclusion, the storage stability of Beriplast P was demonstrated over a range of 24-month storage schedules including interim exposure to elevated temperature, and the reconstituted product was stable for up to 50 h.

  10. Platelet rich fibrin and xenograft in treatment of intrabony defect

    Directory of Open Access Journals (Sweden)

    Saurav Panda

    2014-01-01

    Full Text Available For complete periodontal regeneration, delivery of growth factors in the local environment holds a great deal in adjunct to bone grafts. Platelet rich fibrin (PRF is considered as second generation platelet concentrate, consisting of viable platelets, releasing various growth factors such as platelet-derived growth factor, vascular endothelial growth factor, transforming growth factor, insulin-like growth factor, epidermal growth factor and basic fibroblast growth factor. Hence, this case report aims to investigate the clinical and radiological (bone fill effectiveness of autologous PRF along with the use of xenogenic bone mineral in the treatment of intra bony defects. Intrabony defect was treated with autologous PRF along with the use of xenogenic bone mineral. A decrease in probing pocket depth, gain in clinical attachment level and significant bone fill was observed at end of 6 months. The result obtained with the use of PRF may be attributed to the sustained and simultaneous release of various growth factors over a period of 7 days. In this case report, the positive clinical impact of additional application of PRF with xenogenic graft material in the treatment of periodontal intrabony defect was seen.

  11. Autologous blood preparations rich in platelets, fibrin and growth factors.

    Science.gov (United States)

    Fioravanti, C; Frustaci, I; Armellin, E; Condò, R; Arcuri, C; Cerroni, L

    2015-01-01

    Bone regeneration is often needed prior to dental implant treatment due to the lack of adequate quantity and quality after infectious diseases. The greatest regenerative power was obtained with autologous tissue, primarily the bone alive, taken from the same site or adjacent sites, up to the use centrifugation of blood with the selection of the parts with the greatest potential regenerative. In fact, various techniques and technologies were chronologically successive to cope with an ever better preparation of these concentrates of blood. Our aim is to review these advances and discuss the ways in which platelet concentrates may provide such unexpected beneficial therapeutic effects. The research has been carried out in the MEDLINE and Cochrane Central Register of Controlled Trials database by choosing keywords as "platelet rich plasma", "platelet rich fibrin", "platelet growth factors", and "bone regeneration" and "dentistry". Autologous platelet rich plasma is a safe and low cost procedure to deliver growth factors for bone and soft tissue healing. The great heterogeneity of clinical outcomes can be explained by the different PRP products with qualitative and quantitative difference among substance.

  12. Effect of Platelet-Rich Fibrin on Peripheral Nerve Regeneration.

    Science.gov (United States)

    Şenses, Fatma; Önder, Mustafa E; Koçyiğit, Ismail D; Kul, Oğuz; Aydin, Gülümser; Inal, Elem; Atil, Fethi; Tekin, Umut

    2016-10-01

    This study aimed to evaluate the effect of platelet-rich fibrin (PRF) on peripheral nerve regeneration on the sciatic nerve of rats by using functional, histopathologic, and electrophysiologic analyses. Thirty female Wistar rats were divided randomly into 3 experimental groups. In group 1 (G1), which was the control group, the sciatic nerve was transected and sutured (n = 10). In group 2 (G2), the sciatic nerve was transected, sutured, and then covered with PRF as a membrane (n = 10). In group 3 (G3), the sciatic nerve was transected, sutured by leaving a 5-mm gap, and then covered by PRF as a nerve guide (n = 10). Functional, histopathologic, and electrophysiologic analyses were performed. The total histopathologic semiquantitative score was significantly higher in G1 compared to G2 and G3 (P < 0.05). Myelin thickness and capillaries were significantly lower in G3 compared to G1 (P < 0.05). There was no statistically significant difference between the groups with regard to the functional and electrophysiologic results. The study results suggest that PRF decreases functional recovery in sciatic nerve injury. Further studies are required to determine the efficacy of PRF on peripheral nerve regeneration.

  13. Comparison between electrocautery and fibrin selant after hepatectomy in rats.

    Science.gov (United States)

    Coutinho, Tâmara Rúbia Cavalcante Guimarães; Malafaia, Osvaldo; Torres, Orlando Jorge Martins; Ribas Filho, Jurandir Marcondes; Kaminski, Alvaro Fonseca; Cella, Igor Furlan; Jurkonis, Leandro Bressianini

    2014-01-01

    To compare between electrocautery and fibrin sealant hemostasis in rats after partial hepatectomy. we used 24 Wistar rats, which were submitted to 30% hepatic resection, divided into two groups of 12 animals each: Group Electrocautery and Group Tachosil(r). These animals were evaluated after three and 14 days. We assessed the presence of complications, laboratory tests and histological exam of the recovered liver. the presence of abscess was more prevalent in the electrocautery group. The observed adhesions were more pronounced in the electrocautery group, both in frequency and in intensity, after three and 14 days. There were no deaths in either group. As for laboratory analysis, after three days the hematocrit was lower in the TachoSil(r) Group. The elevation of AST and ALT were more pronounced in the electrocautery group (p = 0.002 and p = 0.004) in three days. Histological analysis of specimens collected on the third day after surgery showed similar results in both groups for the presence of polymorphonuclear cells, whereas mononuclear was more evident in the TachoSil(r) group. We also observed that angiogenesis, although present in both groups, was more pronounced in the TachoSil(r) group (p = 0.030). However, on the 14th day angiogenesis was more pronounced in the electrocautery group, but without statistical significance. hemostasis achieved by the groups was similar; however, the use of electrocautery was associated with infections, adhesions at higher grades and elevated liver enzymes.

  14. Significance of soluble growth factors in the chondrogenic response of human umbilical cord matrix stem cells in a porous three dimensional scaffold

    Directory of Open Access Journals (Sweden)

    RS Nirmal

    2013-11-01

    Full Text Available Stem cell based tissue engineering has emerged as a promising strategy for articular cartilage regeneration. Foetal derived mesenchymal stem cells (MSCs with their ease of availability, pluripotency and high expansion potential have been demonstrated to be an attractive cell source over adult MSCs. However, there is a need for optimisation of chondrogenic signals to direct the differentiation of these multipotent MSCs to chondrogenic lineage. In this study we have demonstrated the in vitro chondrogenesis of human umbilical cord matrix MSCs in three dimensional PVA-PCL (polyvinyl alcohol-polycaprolactone scaffolds in the presence of the individual growth factors TGFβ1, TGFβ3, IGF, BMP2 and their combination with BMP2. Gene expression, histology and immunohistology were evaluated after 28 d culture. The induced cells showed the feature of chondrocytes in their morphology and expression of typical chondrogenic extracellular matrix molecules. Moreover, the real-time PCR assay has shown the expression of gene markers of chondrogenesis, SOX9, collagen type II and aggrecan. The expression of collagen type I and collagen type X was also evaluated. This study has demonstrated the successful chondrogenic induction of human umbilical cord MSCs in 3D scaffolds. Interestingly, the growth factor combination of TGF-β3 and BMP-2 was found to be more effective for chondrogenesis as shown by the real-time PCR studies. The findings of this study suggest the importance of using growth factor combinations for successful chondrogenic differentiation of umbilical cord MSCs.

  15. The immunodominant influenza matrix t cell epitope recognized in human induces influenza protection in HLA-A2/Kb transgenic mice

    International Nuclear Information System (INIS)

    Plotnicky, H.; Cyblat-Chanal, D.; Aubry, J.-P.; Derouet, F.; Klinguer-Hamour, C.; Beck, A.; Bonnefoy, J.-Y.; Corvaiea, N.

    2003-01-01

    The protective efficacy of the influenza matrix protein epitope 58-66 (called M1), recognized in the context of human HLA-A2 molecules, was evaluated in a HLA-A2/K b transgenic mouse model of lethal influenza infection. Repeated subcutaneous immunizations with M1 increased the percentage of survival. This effect was mediated by T cells since protection was abolished following in vivo depletion of all T lymphocytes, CD8 + , or CD4 + T cells. The survival correlated with the detection of memory CD8 + splenocytes able to proliferate in vitro upon stimulation with M1 and to bind M1-loaded HLA-A2 dimers, as well as with M1-specific T cells in the lungs, which were directly cytotoxic to influenza-infected cells following influenza challenge. These results demonstrated for the first time that HLA-A2-restricted cytotoxic T cells specific for the major immunodominant influenza matrix epitope are protective against the infection. They encourage further in vivo evaluation of T cell epitopes recognized in the context of human MHC molecules

  16. Fibrin glue as a protective tool for microanastomoses in limb reconstructive surgery

    Directory of Open Access Journals (Sweden)

    Langer, Stefan

    2015-12-01

    Full Text Available Aim: Fibrin glue becomes a more and more routinely used tool for stabilization of microanastomoses and nerve repair. This paper summarizes the technical properties and advantages of its use in a wide variety of microsurgical contexts, and includes an exemplary limb reconstructive case.Patients and methods: A total of 131 patients who had undergone elective and emergency microsurgery mainly of the limbs were retrospectively analyzed, as was the use of free flaps.Results: The use of fibrin glue allows for proper positioning of anastomoses and repaired nerves. No torsion of the pedicle could be seen. The flap survival rated >94%. The fibrin glue could stay in place in >99%. In the rare case of revision, the fibrin glue could easily be removed without damaging the region of the microanastomosis.Conclusion: Fibrin glue should not be used to repair insufficient, i.e., leaking anastomoses, but it does protect the site of anastomosis from tissue and fluid pressure. It prevents the pedickle from torsion and its use facilitates relocation of the microanastomoses in cases of revision surgery.

  17. Topical tissue plasminogen activator appears ineffective for the clearance of intraocular fibrin.

    Science.gov (United States)

    Zwaan, J; Latimer, W B

    1998-06-01

    To determine the efficacy of topical tissue plasminogen activator (tPA) for the resolution of postoperative or inflammatory intraocular fibrinous exudates. Each treatment consisted of drops of 1 mg/ml tPA given 9 times 5 minutes apart. Records were reviewed and the results at 24 and 48 hours were recorded. Sixty-two patients had a total of 94 treatments. Fibrin exudates following intraocular surgery in 34 patients were treated 44 times. In 6 patients there was a positive result. Fibrin associated with intraocular infection was treated in 9 patients. None showed clear improvement. Nineteen patients had a total of 34 treatments for poorly controlled intraocular pressure (IOP) after glaucoma surgery. Five patients showed adequate control of the IOP, 12 did not change, and 2 had a questionable improvement. Eleven patients had adequate IOP control after additional treatment. Seven required suture lysis, 2 ab interno bleb revision, and 2 YAG capsulotomy or iridotomy to reduce the IOP to an acceptable level. Within the limits of this retrospective study and taking into account that fibrin may resolve spontaneously, it appears that topical tPA drops are not effective for the liquefaction of intraocular fibrin after surgery or in association with intraocular inflammation. They did not improve IOP control after glaucoma surgery.

  18. The application of large amplitude oscillatory stress in a study of fully formed fibrin clots

    Science.gov (United States)

    Lamer, T. F.; Thomas, B. R.; Curtis, D. J.; Badiei, N.; Williams, P. R.; Hawkins, K.

    2017-12-01

    The suitability of controlled stress large amplitude oscillatory shear (LAOStress) for the characterisation of the nonlinear viscoelastic properties of fully formed fibrin clots is investigated. Capturing the rich nonlinear viscoelastic behaviour of the fibrin network is important for understanding the structural behaviour of clots formed in blood vessels which are exposed to a wide range of shear stresses. We report, for the first time, that artefacts due to ringing exist in both the sample stress and strain waveforms of a LAOStress measurement which will lead to errors in the calculation of nonlinear viscoelastic properties. The process of smoothing the waveforms eliminates these artefacts whilst retaining essential rheological information. Furthermore, we demonstrate the potential of LAOStress for characterising the nonlinear viscoelastic properties of fibrin clots in response to incremental increases of applied stress up to the point of fracture. Alternating LAOStress and small amplitude oscillatory shear measurements provide detailed information of reversible and irreversible structural changes of the fibrin clot as a consequence of elevated levels of stress. We relate these findings to previous studies involving large scale deformations of fibrin clots. The LAOStress technique may provide useful information to help understand why some blood clots formed in vessels are stable (such as in deep vein thrombosis) and others break off (leading to a life threatening pulmonary embolism).

  19. Autologous fibrin sealant (Vivostat®) in the neurosurgical practice: Part I: Intracranial surgical procedure

    Science.gov (United States)

    Graziano, Francesca; Certo, Francesco; Basile, Luigi; Maugeri, Rosario; Grasso, Giovanni; Meccio, Flavia; Ganau, Mario; Iacopino, Domenico G.

    2015-01-01

    Background: Hemorrhages, cerebrospinal fluid (CSF) fistula and infections are the most challenging postoperative complications in Neurosurgery. In this study, we report our preliminary results using a fully autologous fibrin sealant agent, the Vivostat® system, in achieving hemostasis and CSF leakage repair during cranio-cerebral procedures. Methods: From January 2012 to March 2014, 77 patients were studied prospectively and data were collected and analyzed. Autologous fibrin sealant, taken from patient's blood, was prepared with the Vivostat® system and applied on the resection bed or above the dura mater to achieve hemostasis and dural sealing. The surgical technique, time to bleeding control and associated complications were recorded. Results: A total of 79 neurosurgical procedures have been performed on 77 patients. In the majority of cases (98%) the same autologous fibrin glue provided rapid hemostasis and dural sealing. No patient developed allergic reactions or systemic complications in association with its application. There were no cases of cerebral hematoma, swelling, infection, or epileptic seizures after surgery whether in the immediate or in late period follow-up. Conclusions: In this preliminary study, the easy and direct application of autologous fibrin sealant agent helped in controlling cerebral bleeding and in providing prompt and efficient dural sealing with resolution of CSF leaks. Although the use of autologous fibrin glue seems to be safe, easy, and effective, further investigations are strongly recommended to quantify real advantages and potential limitations. PMID:25984391

  20. Efficacy of fibrin-sealants in reducing biliary leakage following laparoscopic common bile duct exploration.

    Science.gov (United States)

    Parra-Membrives, Pablo; Martínez-Baena, Darío; Lorente-Herce, José Manuel; Martín-Balbuena, Ramón

    2018-05-21

    In spite of the acquired experience with laparoscopic common bile duct exploration (LCBDE) for choledocholithiasis management, there is still a risk of biliary leakage of 5% to 15% following choledochotomy closure. We evaluate the usefulness of fibrin-collagen sealants to reduce the incidence of biliary fistula after laparoscopic choledochorrhaphy. We report a retrospective analysis of 96 patients undergoing LCBDE from March 2009 to March 2017, whose closure of the bile duct was completed by antegrade stenting and choledochorraphy or by performing a primary suture. The study population was divided into two groups according to whether they received a collagen-fibrin sealant covering the choledochorrhaphy or not, analyzing the incidence of postoperative biliary fistula in each group. Thirty-nine patients (41%) received a fibrin-collagen sponge while the bile duct closure was not covered in the remaining 57 patients (59%). The incidence of biliary fistula was 7.7% (3 patients) in the first group and 14% (8 patients) in the second group (P=.338). In patients who underwent primary choledochorraphy, the fibrin-collagen sealant reduced the incidence of biliary leakage significantly (4.5% vs. 33%, P=.020), which was a protective factor with an odds ratio of 10.5. Fibrin-collagen sealants may decrease the incidence of biliary fistula in patients who have undergone primary bile duct closure following LCBDE. Copyright © 2018 AEC. Publicado por Elsevier España, S.L.U. All rights reserved.

  1. Excess Fibrin Deposits Decrease Fetal Weight of Pregnant Mice Infected by Plasmodium berghei

    Directory of Open Access Journals (Sweden)

    Desy Andari

    2014-05-01

    Full Text Available Low birth weight is commonly attributed to malaria in pregnancy, but the cellular and molecular mechanisms that underlie this poor birth outcome are incompletely understood. A universally described histopathological feature of placental malaria is excessive deposition of fibrin, the end-product of the coagulation cascade. This study was conducted to compare fibrin deposit in pregnant mice that infected by Plasmodium berghei (treatment group to the normal pregnant mice (control group and its association with fetal weight. This research is in vivo experimental laboratory study that used 18 pregnant Balb/c mice which divided to the control the group (8 mice and treatment group (9 mice infected by P.berghei. Placentas were staining with Haematoxylin-Eosin (HE for fibrin deposits examination whereas fetal weight was performed with Mettler analytical balance AE 50. Fetal weight of the treatment group was lower than those of the control group (t test, p=0,002. Fibrin deposits were increased in the treatment group (t test, p=0,005 and influenced weight of fetuses (Spearman r= -0,586, p= 0,014. Weights of fetuses are interfered by fibrin deposits during malaria infection.

  2. Characterization of Fibrin and Collagen Gels for Engineering Wound Healing Models

    Directory of Open Access Journals (Sweden)

    Oihana Moreno-Arotzena

    2015-04-01

    Full Text Available Hydrogels are used for 3D in vitro assays and tissue engineering and regeneration purposes. For a thorough interpretation of this technology, an integral biomechanical characterization of the materials is required. In this work, we characterize the mechanical and functional behavior of two specific hydrogels that play critical roles in wound healing, collagen and fibrin. A coherent and complementary characterization was performed using a generalized and standard composition of each hydrogel and a combination of techniques. Microstructural analysis was performed by scanning electron microscopy and confocal reflection imaging. Permeability was measured using a microfluidic-based experimental set-up, and mechanical responses were analyzed by rheology. We measured a pore size of 2.84 and 1.69 μm for collagen and fibrin, respectively. Correspondingly, the permeability of the gels was 1.00·10−12 and 5.73·10−13 m2. The shear modulus in the linear viscoelastic regime was 15 Pa for collagen and 300 Pa for fibrin. The gels exhibited strain-hardening behavior at ca. 10% and 50% strain for fibrin and collagen, respectively. This consistent biomechanical characterization provides a detailed and robust starting point for different 3D in vitro bioapplications, such as collagen and/or fibrin gels. These features may have major implications for 3D cellular behavior by inducing divergent microenvironmental cues.

  3. Anatomical region-dependent enhancement of 3-dimensional chondrogenic differentiation of human mesenchymal stem cells by soluble meniscus extracellular matrix.

    Science.gov (United States)

    Rothrauff, Benjamin B; Shimomura, Kazunori; Gottardi, Riccardo; Alexander, Peter G; Tuan, Rocky S

    2017-02-01

    Extracellular matrix (ECM) derived from decellularized tissues has been found to promote tissue neogenesis, most likely mediated by specific biochemical and physical signaling motifs that promote tissue-specific differentiation of progenitor cells. Decellularized ECM has been suggested to be efficacious for the repair of tissue injuries. However, decellularized meniscus contains a dense collagenous structure, which impedes cell seeding and infiltration and is not readily applicable for meniscus repair. In addition, the meniscus consists of two distinct anatomical regions that differ in vascularity and cellular phenotype. The purpose of this study was to explore the region-specific bioactivity of solubilized ECM derived from the inner and outer meniscal regions as determined in 2D and 3D cultures of adult mesenchymal stem cells (MSCs). When added as a medium supplement to 2D cultures of MSCs, urea-extracted fractions of the inner (imECM) and outer meniscal ECM (omECM) enhanced cell proliferation while imECM most strongly upregulated fibrochondrogenic differentiation on the basis of gene expression profiles. When added to 3D cultures of MSCs seeded in photocrosslinked methacrylated gelatin (GelMA) hydrogels, both ECM fractions upregulated chondrogenic differentiation as determined by gene expression and protein analyses, as well as elevated sulfated glycosaminoglycan sGAG content, compared to ECM-free controls. The chondrogenic effect at day 21 was most pronounced with imECM supplementation, but equivalent between ECM groups by day 42. Despite increased cartilage matrix, imECM and omECM constructs possessed compressive moduli similar to controls. In conclusion, soluble meniscal ECM may be considered for use as a tissue-specific reagent to enhance chondrogenesis for MSC-based 3D cartilage tissue engineering. The inner region of the knee meniscus is frequently injured and possesses a poor intrinsic healing capacity. Solubilized extracellular matrix (ECM) derived from

  4. Matrix Metalloproteinase-9/Neutrophil Gelatinase-Associated Lipocalin Complex Activity in Human Glioma Samples Predicts Tumor Presence and Clinical Prognosis

    Directory of Open Access Journals (Sweden)

    Ming-Fa Liu

    2015-01-01

    Full Text Available Matrix metalloproteinase-9/neutrophil gelatinase-associated lipocalin (MMP-9/NGAL complex activity is elevated in brain tumors and may serve as a molecular marker for brain tumors. However, the relationship between MMP-9/NGAL activity in brain tumors and patient prognosis and treatment response remains unclear. Here, we compared the clinical characteristics of glioma patients with the MMP-9/NGAL activity measured in their respective tumor and urine samples. Using gelatin zymography assays, we found that MMP-9/NGAL activity was significantly increased in tumor tissues (TT and preoperative urine samples (Preop-1d urine. Activity was reduced by seven days after surgery (Postop-1w urine and elevated again in cases of tumor recurrence. The MMP-9/NGAL status correlated well with MRI-based tumor assessments. These findings suggest that MMP-9/NGAL activity could be a novel marker to detect gliomas and predict the clinical outcome of patients.

  5. Cooperative effects in differentiation and proliferation between PDGF-BB and matrix derived synthetic peptides in human osteoblasts

    Directory of Open Access Journals (Sweden)

    Vordemvenne Thomas

    2011-11-01

    Full Text Available Abstract Background Enhancing osteogenic capabilities of bone matrix for the treatment of fractures and segmental defects using growth factors is an active area of research. Recently, synthetic peptides like AC- 100, TP508 or p-15 corresponding to biologically active sequences of matrix proteins have been proven to stimulate bone formation. The platelet-derived growth factor (PDGF BB has been identified as an important paracrine factor in early bone healing. We hypothesized that the combined use of PDGF-BB with synthetic peptides could result in an increase in proliferation and calcification of osteoblast-like cells. Methods Osteoblast-like cell cultures were treated with PDGF and synthetic peptides, singly and as combinations, and compared to non-treated control cell cultures. The cultures were evaluated at days 2, 5, and 10 in terms of cell proliferation, calcification and gene expression of alkaline phosphate, collagen I and osteocalcin. Results Experimental findings revealed that the addition of PDGF, p-15 and TP508 and combinations of PDGF/AC-100, PDGF/p-15 and PDGF/TP508 resulted in an increase in proliferating osteoblasts, especially in the first 5 days of cultivation. Proliferation did not significantly differ between single factors and factor combinations (p > 0.05. The onset of calcification in osteoblasts occurred earlier and was more distinct compared to the corresponding control or PDGF stimulation alone. Significant difference was found for the combined use of PDGF/p-15 and PDGF/AC-100 (p Conclusions Our findings indicate that PDGF exhibits cooperative effects with synthetic peptides in differentiation and proliferation. These cooperative effects cause a significant early calcification of osteoblast-like cells (p

  6. Crosslinking of fihrinogen by factor XIHa exposes fibrin-specific epitopes and induces enhanced plasminogen activation by t-PA

    NARCIS (Netherlands)

    Nieuwenhuizen, W.; Voskuilen, M.; Kevin, R.; Siebenlis; Michael, W.; Mosesson

    1996-01-01

    'Fibrin-specific'epitopes 'Aαl48-160' and 'γ312-324' are exposed when fibrinogen (Fbg) is converted to fibrin (Fb). Particularly, polymerized Fb enhances the t-PA-mediated plasminogen activation. Fb polymerization involves 'D:E' assembly, end-to-end self-association ('D:D') and interactions between

  7. Successful Endoscopic Management of Non-Healing Perforated Duodenal Ulcer with Polyglycolic Acid Sheet and Fibrin Glue.

    Science.gov (United States)

    Mishiro, Tsuyoshi; Shibagaki, Kotaro; Matsuda, Kayo; Fukuyama, Chika; Okada, Mayumi; Mikami, Hironobu; Izumi, Daisuke; Yamashita, Noritsugu; Okimoto, Eiko; Fukuda, Naoki; Aimi, Masahito; Fukuba, Nobuhiko; Oshima, Naoki; Takanashi, Toshihiro; Matsubara, Takeshi; Ishimura, Norihisa; Ishihara, Shunji; Kinoshita, Yoshikazu

    2016-08-01

    In recent years, treatment techniques in which polyglycolic acid sheets are applied to various situations with fibrin glue have exhibited great clinical potential, and previous studies have reported safety and efficacy. We describe closure of a non-healing perforated duodenal ulcer with the use of a polyglycolic acid sheet and fibrin glue in an elderly patient who was not a candidate for surgery.

  8. Fibrin hydrogels to deliver dental stem cells of the apical papilla for regenerative medicine.

    Science.gov (United States)

    Germain, Loïc; De Berdt, Pauline; Vanacker, Julie; Leprince, Julian; Diogenes, Anibal; Jacobs, Damien; Vandermeulen, Gaëlle; Bouzin, Caroline; Préat, Véronique; Dupont-Gillain, Christine; des Rieux, Anne

    2015-01-01

    Evaluation of survival, proliferation and neurodifferentiation of dental stem cells from the apical papilla (SCAP) in fibrin hydrogels. We hypothesized that fibrin composition will influence cell behavior. Modulus, pore and fiber size were measured. SCAP in vitro viability, proliferation and neural differentiation, as well as in vivo proliferation and angiogenesis were studied. Hydrogel moduli were influenced by fibrin formulation but not hydrogel morphology, SCAP in vitro viability and proliferation. In total 60% of SCAP expressed PanNeurofilament in vitro without induction in Fibrinogen50-Thrombin10. SCAP proliferated when implanted in vivo and stimulated host endothelial cell infiltration. Fibrinogen30-Thrombin10 or Thrombin50 would be more favorable to in vitro SCAP viability and in vivo proliferation, while Fibrinogen 50-Thrombin50 would be more adapted to neurodifferentiation.

  9. Platelet-rich fibrin in the treatment of periodontal bone defects.

    Science.gov (United States)

    Ranganathan, Aravindhan T; Chandran, Chitraa R

    2014-05-01

    Periodontitis is characterized by the formation of true pockets, bone loss and attachment loss. Various techniques have been attempted in the past to truly regenerate the lost periodontal structures, albeit with variable outcome. In this evolution, the technique being tried out widely is the use of platelet rich concentrates, namely platelet-rich fibrin (PRF). In this report, we present a case of surgical treatment of osseous bone defects namely two walled crater and dehiscence treated in posterior teeth with autologously prepared platelet rich fibrin mixed with hydroxy apatite bone graft and PRF in the form of a membrane. Our results showed clinical improvements in all the clinical parameters postoperatively namely the pocket depth reduction and gain in attachment level and hence, PRF can be used alone or in combination with the bone graft to yield successful clinical results in treating periodontal osseous defects. Platelet-rich fibrin is an effective alternative to platelet-rich plasma (PRP) in reconstructing bone defects.

  10. Developing Mesoscale Model of Fibrin-Platelet Network Representing Blood Clotting =

    Science.gov (United States)

    Sun, Yueyi; Nikolov, Svetoslav; Bowie, Sam; Alexeev, Alexander; Lam, Wilbur; Myers, David

    Blood clotting disorders which prevent the body's natural ability to achieve hemostasis can lead to a variety of life threatening conditions such as, excessive bleeding, stroke, or heart attack. Treatment of these disorders is highly dependent on understanding the underlying physics behind the clotting process. Since clotting is a highly complex multi scale mechanism developing a fully atomistic model is currently not possible. We develop a mesoscale model based on dissipative particle dynamics (DPD) to gain fundamental understanding of the underlying principles controlling the clotting process. In our study, we examine experimental data on clot contraction using stacks of confocal microscopy images to estimate the crosslink density in the fibrin networks and platelet location. Using this data we reconstruct the platelet rich fibrin network and study how platelet-fibrin interactions affect clotting. Furthermore, we probe how different system parameters affect clot contraction. ANSF CAREER Award DMR-1255288.

  11. Clinical experience of intrapleural administration of fibrin glue for secondary pneumothorax with advanced lung c