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1
Hamati, H. F., E. L. Britton, and D. J. Carey. "Inhibition of proteoglycan synthesis alters extracellular matrix deposition, proliferation, and cytoskeletal organization of rat aortic smooth muscle cells in culture." Journal of Cell Biology 108, no. 6 (June 1, 1989): 2495–505. http://dx.doi.org/10.1083/jcb.108.6.2495.
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Arterial proteoglycans have been implicated in several important physiological processes ranging from lipid metabolism to regulation of smooth muscle cell growth. Vascular smooth muscle (VSM) cells are the major producers of proteoglycans in the medial layer of blood vessels. To study functional consequences of alterations in VSM proteoglycan metabolism we used 4-methylumbelliferyl-beta-D-xyloside to inhibit proteoglycan synthesis in primary and early passage cultures of rat aortic smooth muscle cells. Biochemical analysis of cultures labeled with 35SO4 showed the drug inhibited synthesis of different classes of proteoglycans by 50 to 62%. Inhibition of proteoglycan synthesis resulted in reduced accumulation of extracellular matrix, as shown by immunofluorescent staining with antibodies to chondroitin sulfate, fibronectin, thrombospondin, and laminin. There was also an inhibition of postconfluent (multilayered) growth of the smooth muscle cells, and a change in the morphology of the cells, with no apparent effect on subconfluent growth. In addition, in drug-treated cells there was a reduction in the number of cytoskeletal filaments that contained alpha-actin, the actin subtype synthesized by differentiated VSM cells. This occurred even though the total content of alpha-actin in the cells was not reduced. The effects of the inhibitor on growth and morphology could be reversed by switching the cultures to normal medium and could be prevented by growing the cells on preformed VSM extracellular matrix. These observations suggest the vascular extracellular matrix may play a role in regulating the growth and differentiation of smooth muscle cells.
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Sah, R. L. Y., A. J. Grodzinsky, A. H. K. Plaas, and J. D. Sandy. "Effects of tissue compression on the hyaluronate-binding properties of newly synthesized proteoglycans in cartilage explants." Biochemical Journal 267, no. 3 (May 1, 1990): 803–8. http://dx.doi.org/10.1042/bj2670803.
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The effects of tissue compression on the hyaluronate-binding properties of newly synthesized proteoglycans in calf cartilage explants were examined. Pulse-chase experiments showed that conversion of low-affinity monomers to the high-affinity form (that is, to a form capable of forming aggregates with 1.6% hyaluronate on Sephacryl S-1000) occurred with a t1/2 of about 5.7 h in free-swelling discs at pH 7.45. Static compression during chase (in pH 7.45 medium) slowed the conversion, as did incubation in acidic medium (without compression). Both effects were dose-dependent. For example, the t1/2 for conversion was increased to about 11 h by either (1) compression from a thickness of 1.25 mm to 0.5 mm or (2) medium acidification from pH 7.45 to 6.99. Oscillatory compression of 2% amplitude at 0.001, 0.01, or 0.1 cycles/s during chase did not, however, affect the conversion. Changes in the hyaluronate-binding affinity of [35S]proteoglycans in these experiments were accompanied by no marked change in the high percentage (approximately 80%) of monomers which could form aggregates with excess hyaluronate and link protein. Since static tissue compression would result in an increased matrix proteoglycan concentration and thereby a lower intra-tissue pH [Gray, Pizzanelli, Grodzinsky & Lee (1988) J. Orthop. Res. 6, 777-792], it seems likely that matrix pH may influence proteoglycan aggregate assembly by an effect on the hyaluronate-binding affinity of proteoglycan monomer. Such a pH mechanism might have a physiological role, promoting proteoglycan deposition in regions of low proteoglycan concentration.
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Kolb, Martin, Peter J. Margetts, Patricia J. Sime, and Jack Gauldie. "Proteoglycans decorin and biglycan differentially modulate TGF-β-mediated fibrotic responses in the lung." American Journal of Physiology-Lung Cellular and Molecular Physiology 280, no. 6 (June 1, 2001): L1327—L1334. http://dx.doi.org/10.1152/ajplung.2001.280.6.l1327.
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Transforming growth factor (TGF)-β is a key cytokine in the pathogenesis of pulmonary fibrosis, and pharmacological interference with TGF-β can ameliorate the fibrotic tissue response. The small proteoglycans decorin and biglycan are able to bind and inhibit TGF-β activity in vitro. Although decorin has anti-TGF-β properties in vivo, little is known about the physiological role of biglycan in vivo. Adenoviral gene transfer was used to overexpress active TGF-β, decorin, and biglycan in cell culture and in murine lungs. Both proteoglycans were able to interfere with TGF-β bioactivity in vitro in a dose-dependant manner. In vivo, overexpression of TGF-β resulted in marked lung fibrosis, which was significantly reduced by concomitant overexpression of decorin. Biglycan, however, had no significant effect on lung fibrosis induced by TGF-β. The data suggest that differences in tissue distribution are responsible for the different effects on TGF-β bioactivity in vivo, indicating that decorin, but not biglycan, has potential therapeutic value in fibrotic disorders of the lung.
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Schröer, Katrin, Montaha Alshawabkeh, Sebastian Schellhorn, Katrin Bronder, Wenli Zhang, and Anja Ehrhardt. "Influence of Heparan Sulfate Proteoglycans and Factor X on species D Human Adenovirus Uptake and Transduction." Viruses 15, no. 1 (December 24, 2022): 55. http://dx.doi.org/10.3390/v15010055.
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More than 100 human adenovirus (Ad) types were identified, of which species D comprises the largest group. Heparan sulfate proteoglycans (HSPGs) were shown to function as cell surface receptors for cell binding and uptake of some Ads, but a systematic analysis of species D Ads is lacking. Previous research focused on Ad5 and blood coagulation factor X (FX) complexes, which revealed that Ad5 can transduce cells with low expression levels of its main coxsackievirus-adenovirus receptor in the presence of high HSPG expression levels in a FX dependent manner. Based on our reporter gene-tagged Ad-library, we explored for the first time a broad spectrum of species D Ads to study the role of HSPG on their cellular uptake. This study was performed on three Chinese Hamster Ovary (CHO) cell lines with different forms of HSPG (only proteoglycan (745), non-sulfated HSPG (606) or sulfated HSPG (K1)). The effect of Ad:FX complexes on Ad uptake was explored in the presence of physiological levels of FX in blood (6–10 µg/mL). We found that sulfation of HSPG plays an important role in cellular uptake and transduction of FX-bound Ad5 but neither HSPG nor FX influenced uptake of all tested species D Ads. Because FX has no influence on transduction efficiencies of species D Ads and therefore may not bind to them, these Ads may not be protected from attack by neutralizing IgM antibodies or the complement pathway, which may have implications for species D Ads used as vaccine and gene therapy vectors.
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Váncza, Lórand, Péter Tátrai, Andrea Reszegi, Kornélia Baghy, and Ilona Kovalszky. "SPOCK1 with unexpected function. The start of a new career." American Journal of Physiology-Cell Physiology 322, no. 4 (April 1, 2022): C688—C693. http://dx.doi.org/10.1152/ajpcell.00033.2022.
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SPOCK1, 2, and 3 are considered matricellular proteoglycans without a structural role. Their functions are only partly elucidated. SPOCK1 was detected in the brain as a member of the neural synapses, then in the neuromuscular junctions. It plays a role in the regulation of the blood-brain barrier. Its best-characterized activity was its oncogenic potential discovered in 2012. Its deleterious effect on tumor progression was detected on 36 different types of tumors by the end of 2020. However, its mode of action is still not completely understood. Furthermore, even less was discovered about its physiological function. The fact that it was found to localize in the mitochondria and interfered with the lipid metabolism indicated that the full discovery of SPOCK1 is still waiting for us.
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Al-Jamal, Rehab, and Mara S. Ludwig. "Changes in proteoglycans and lung tissue mechanics during excessive mechanical ventilation in rats." American Journal of Physiology-Lung Cellular and Molecular Physiology 281, no. 5 (November 1, 2001): L1078—L1087. http://dx.doi.org/10.1152/ajplung.2001.281.5.l1078.
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Excessive mechanical ventilation results in changes in lung tissue mechanics. We hypothesized that changes in tissue properties might be related to changes in the extracellular matrix component proteoglycans (PGs). The effect of different ventilation regimens on lung tissue mechanics and PGs was examined in an in vivo rat model. Animals were anesthetized, tracheostomized, and ventilated at a tidal volume of 8 (Vt 8), 20, or 30 (Vt 30) ml/kg, positive end-expiratory pressure of 0 (PEEP0) or 1.5 (PEEP1.5) cmH2O, and frequency of 1.5 Hz for 2 h. The constant-phase model was used to derive airway resistance, tissue elastance, and tissue damping. After physiological measurements, one lung was frozen for immunohistochemistry and the other was reserved for PG extraction and Western blotting. After 2 h of mechanical ventilation, tissue elastance and damping were significantly increased in rats ventilated at Vt 30PEEP0 compared with control rats (ventilated at Vt 8PEEP1.5). Versican, basement membrane heparan sulfate PG, and biglycan were all increased in rat lungs ventilated at Vt 30PEEP0 compared with control rats. At Vt 30PEEP0, heparan sulfate PG and versican staining became prominent in the alveolar wall and airspace; biglycan was mostly localized in the airway wall. These data demonstrate that alterations in lung tissue mechanics with excessive mechanical ventilation are accompanied by changes in all classes of extracellular matrix PG.
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Valentim da Silva, Rodrigo Marcel, Priscila Arend Barichello, Melyssa Lima Medeiros, Waléria Cristina Miranda de Mendonça, Jung Siung Camel Dantas, Oscar Ariel Ronzio, Patricia Meyer Froes, and Hassan Galadari. "Effect of Capacitive Radiofrequency on the Fibrosis of Patients with Cellulite." Dermatology Research and Practice 2013 (2013): 1–6. http://dx.doi.org/10.1155/2013/715829.
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Background. Cellulite is a type of lipodystrophy that develops primarily from an alteration in blood circulation or of the lymphatic system that causes structural changes in subcutaneous adipose tissue, collagen, and adjacent proteoglycans. The radiofrequency devices used for cutaneous applications have shown different physiological treatment effects, but there is controversy about the suitable parameters for this type of treatment.Objectives. The aim of this study was to evaluate the effects of low-temperature radiofrequency to confirm the thinning of the collagen tissue and interlobular septa and consequent improvement of cellulite.Methods. A sample of eight women was used to collect ultrasonographic data with a 12 MHz probe that measured collagen fiber thickness. The Vip Electromedicina (Argentina) device, frequency of 0.55 MHz and active electrode 3.5 cm in diameter (area = 9.61 cm2), was applied to a 10 cm2region of the gluteal region for 2 minutes per area of active electrode, during 10 biweekly sessions.Results. The Wilcoxon matched paired test was applied using GraphPad InStat 3.01 for Win95-NT software. Pre- and posttreatment mean collagen fiber thickness showed a 24.66% reduction from 1.01 to 0.67 mm. Statistical analysis using the Wilcoxon matched paired test obtained a significant two-tailedPvalue of 0.0391.Conclusion. It was concluded that the use of more comfortable temperatures favored a reduction in fibrous septum thickness and consequent cellulite improvement, evidenced by the lower degree of severity and decrease in interlobular septal thickness.
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Gilbert, Sophie Jane, Cleo Selina Bonnet, and Emma Jane Blain. "Mechanical Cues: Bidirectional Reciprocity in the Extracellular Matrix Drives Mechano-Signalling in Articular Cartilage." International Journal of Molecular Sciences 22, no. 24 (December 18, 2021): 13595. http://dx.doi.org/10.3390/ijms222413595.
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The composition and organisation of the extracellular matrix (ECM), particularly the pericellular matrix (PCM), in articular cartilage is critical to its biomechanical functionality; the presence of proteoglycans such as aggrecan, entrapped within a type II collagen fibrillar network, confers mechanical resilience underweight-bearing. Furthermore, components of the PCM including type VI collagen, perlecan, small leucine-rich proteoglycans—decorin and biglycan—and fibronectin facilitate the transduction of both biomechanical and biochemical signals to the residing chondrocytes, thereby regulating the process of mechanotransduction in cartilage. In this review, we summarise the literature reporting on the bidirectional reciprocity of the ECM in chondrocyte mechano-signalling and articular cartilage homeostasis. Specifically, we discuss studies that have characterised the response of articular cartilage to mechanical perturbations in the local tissue environment and how the magnitude or type of loading applied elicits cellular behaviours to effect change. In vivo, including transgenic approaches, and in vitro studies have illustrated how physiological loading maintains a homeostatic balance of anabolic and catabolic activities, involving the direct engagement of many PCM molecules in orchestrating this slow but consistent turnover of the cartilage matrix. Furthermore, we document studies characterising how abnormal, non-physiological loading including excessive loading or joint trauma negatively impacts matrix molecule biosynthesis and/or organisation, affecting PCM mechanical properties and reducing the tissue’s ability to withstand load. We present compelling evidence showing that reciprocal engagement of the cells with this altered ECM environment can thus impact tissue homeostasis and, if sustained, can result in cartilage degradation and onset of osteoarthritis pathology. Enhanced dysregulation of PCM/ECM turnover is partially driven by mechanically mediated proteolytic degradation of cartilage ECM components. This generates bioactive breakdown fragments such as fibronectin, biglycan and lumican fragments, which can subsequently activate or inhibit additional signalling pathways including those involved in inflammation. Finally, we discuss how bidirectionality within the ECM is critically important in enabling the chondrocytes to synthesise and release PCM/ECM molecules, growth factors, pro-inflammatory cytokines and proteolytic enzymes, under a specified load, to influence PCM/ECM composition and mechanical properties in cartilage health and disease.
9
Alter, S. C., D. D. Metcalfe, T. R. Bradford, and L. B. Schwartz. "Regulation of human mast cell tryptase. Effects of enzyme concentration, ionic strength and the structure and negative charge density of polysaccharides." Biochemical Journal 248, no. 3 (December 15, 1987): 821–27. http://dx.doi.org/10.1042/bj2480821.
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Tryptase was previously shown to undergo rapid inactivation under physiological conditions unless stabilized by the presence of heparin. The current study shows that increasing the concentration of free tryptase enhances the preservation of enzymic activity, consistent with dissociation of the tetramer, rather than autodegradation, as the mechanism of inactivation. Heparin glycosaminoglycan fragments of Mr greater than 5700 are necessary for complete stabilization of tryptase activity. This stabilizing effect depends upon negative charge density rather than carbohydrate composition. Thus, keratan sulphate or hyaluronic acid were no better than physiological buffer alone; chondroitin monosulphates and heparan sulphate each prolonged the t1/2 about 20-fold over buffer alone; chondroitin sulphate E prolonged the t1/2 69-fold; and dextran sulphate and heparin provided complete stabilization of tryptase activity for 120 min. Poly-D-glutamic acid prolonged the t1/2 55-fold. In each case the loss of tryptase activity followed apparent first-order kinetics. Increasing the NaCl concentration from 0.01 M to 1.0 M increased the stability of free tryptase. In contrast, increasing the NaCl concentration in the presence of stabilizing polysaccharides decreased the stability of tryptase until dissociation of tryptase from each polysaccharide presumably occurred; thereafter tryptase stability increased as did that of free tryptase. The effect of salt concentration on heparin-stabilized tryptase activity (as opposed to stability) was also evaluated. The mast cell proteoglycans heparin and chondroitin sulphate E, by virtue of containing the naturally occurring glycosaminoglycans of highest negative charge density, may play a major role in the regulation of mast cell tryptase activity in vivo.
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Lee, Young Hun, Jun Hyoung Park, Dong Huey Cheon, Taeyoung Kim, Yae Eun Park, Eok-Soo Oh, Ji Eun Lee, and Seung-Taek Lee. "Processing of syndecan-2 by matrix metalloproteinase-14 and effect of its cleavage on VEGF-induced tube formation of HUVECs." Biochemical Journal 474, no. 22 (November 1, 2017): 3719–32. http://dx.doi.org/10.1042/bcj20170340.
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Syndecans (SDCs) are transmembrane proteoglycans that are involved in cell adhesion and cell communication. Specifically, SDC2 plays a key role in tumorigenesis, metastasis, and angiogenesis. Previously, we found that rat SDC2 is shed by matrix metalloproteinase-7 (MMP-7) in colon cancer cells. Here, we analyzed the susceptibility of rat SDC2 to various MMPs. We found that the rat SDC2 ectodomain (ECD) fused to the C-terminal Fc region, which was expressed in mammalian cells, was cleaved more efficiently by MMP-14 than MMP-7. Likewise, when anchored on the surface of HeLa cells, rat SDC2 was cleaved more efficiently by the treatment of MMP-14 than MMP-7 and was shed more readily by membrane-anchored MMP-14 than soluble MMP-14. Furthermore, MMP-14 cleaved recombinant SDC2-ECD expressed in Escherichia coli into multiple fragments. Using N-terminal amino acid sequencing and the top-down proteomics approach, we determined that the major cleavage sites were S88↓L89, T98↓M99, T100↓L101, D132↓P133, and N148↓L149 for rat SDC2-ECD and S55↓G56, S65↓P66, P75↓K76, N92↓I93 D122↓P123, and S138↓L139 for human SDC2-ECD. Finally, the rat and human SDC2-ECD lost the ability to suppress vascular endothelial growth factor-induced formation of capillary-like tubes by human umbilical vein endothelial cells following cleavage by MMP-14, but its major cleavage-site mutant of rat SDC2-ECD did not. These results suggest that MMP-14 is a novel enzyme responsible for degrading SDC2 and impairing its physiological roles including angiogenesis.
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Kim, S. W., M. J. Lee, B. C. Yang, G. S. Im, H. H. Seong, B. S. Yang, H. T. Cheong, and D. H. Kim. "186 THE EFFECT OF MATRIGEL ON THE DEVELOPMENT OF IN VITRO-FERTILIZED PORCINE EMBRYOS." Reproduction, Fertility and Development 19, no. 1 (2007): 209. http://dx.doi.org/10.1071/rdv19n1ab186.
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The application of matrix proteins to culture systems for growth of embryos is a logical extension in the quest to better simulate the in vivo culture environment. Matrigel, a commercially available extracellular matrix product containing collagen IV, laminin, entactin, and proteoglycans isolated from mouse tumor cells, has been tested. Development of mouse pre-implantation embryos cultivated in conventional culture medium was contrasted to that of embryos grown in solubilized Matrigel medium. In the solubilized Matrigel medium, in vitro blastocyst formation and hatching were significantly enhanced over that observed in the medium alone control. Therefore, the aim of this study was to investigate the effect of solubilized Matrigel on the development of porcine embryos after in vitro fertilization. In vitro-matured oocytes were fertilized in mTBM medium with fresh spermatozoa for 6 h. Putative zygotes were cultured in PZM-3 medium supplemented with (matrigel group) or without (control group) 0.8% Matrigel for 6 days. The number of cells in blastocysts was determined by staining with Hoechst 33342. Assessment of apoptosis in blastocysts was examined by TUNEL. The statistical significance of the data was analyzed using chi-square test and Student's t-test. The addition of Matrigel appeared not to increase the proportion of blastocysts (control: 71/219, 21.8 � 2.2% vs. Matrigel: 69/220, 23.5 � 5.8%). However, the mean cell numbers were significantly increased by Matrigel (Matrigel: n = 31, 52.9 � 18.1 vs. control: n = 30, 42.3 � 14.4; P < 0.01). The proportion of apoptotic cells was significantly lower in the Matrigel group (Matrigel: 4.5 � 4.2% vs. control: 6.6 � 5.5%; P < 0.05). In this experiment, Matrigel appeared to increase blastocyst quality of porcine embryos. Results suggest that Matrigel, as an extracellular matrix component, may be another avenue for formulating more physiological culture systems.
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Chen, Yunliang, Michael Scully, Gloria Dawson, Christopher Goodwin, Min Xia, Xinjie Lu, and Ajay Kakkar. "Perturbation of the heparin/heparin-sulfate interactome of human breast cancer cells modulates pro-tumourigenic effects associated with PI3K/Akt and MAPK/ERK signalling." Thrombosis and Haemostasis 109, no. 06 (2013): 1148–57. http://dx.doi.org/10.1160/th12-12-0935.
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SummaryHeparansulfate-proteoglycans (HSPGs) interact via their polyanionic heparansulfate (HS) side chains with a variety of proteins on the cell surface or within the extracellular matrix membrane. The large number of heparin/HS binding proteins form a highly interconnected functional network, which has been termed as the heparin/HS interactome and is functionally linked to physiological and pathological processes. The aim of this study was to investigate the global effect of these protein-HSPG interactions on the tumourigenicity of two breast cancer cell lines (MCF-7 and MDA-MB-231). Cancer cells were cultured in serum-free medium and treated with a concentration of heparin which was capable of modulating HS/ligand interaction. Microarray analysis of MCF-7 cells cultured under these conditions showed that expression of 105 of 1,357 genes potentially related to the pathogenesis of breast neoplasm was significantly altered by heparin treatment. The changes in gene expression correlated with a less tumourigenic phenotype, including reduction of cell adhesive, invasive and migratory properties. These effects were associated with an inhibition of the PI3K/Akt and Raf/MEK/ERK signalling pathways. The modulatory effect of heparin on HS-associated activity was confirmed with one example of heparin/HS interactomes, transforming growth factor β (TGFβ). The innate TGFβ activity of MCF-7 cells was reduced by heparin treatment, with specific interruption of the TGFβ–Smad signalling pathway. The pro-tumourigenic contribution of the heparin/HS interactomes was verified in cells in which HSPG synthesis was blocked using β-xyloside. In conclusion, the interaction between cell surface HPSGs and innate heparin/HS interactomes makes a significant contribution to the tumourigenicity.
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Wan, Lu Ming, Shi Kun Zhang, Su Bo Li, Wen Li, Shou Ping Ji, Lin Gong, Zhi Min Yun, et al. "Heparanase Facilitates PMA-Induced Megakaryocytic Differentiation in K562 Cells via Interleukin 6/STAT3 Pathway." Thrombosis and Haemostasis 120, no. 04 (April 2020): 647–57. http://dx.doi.org/10.1055/s-0040-1705117.
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AbstractHeparanase (HPSE) is an endo-β-D-glucuronidase that cleaves heparan sulfate and hence participates in remodeling of the extracellular matrix, leading to release of cytokines that are immobilized by binding to heparan sulfate proteoglycans (HSPGs), and consequently activating signaling pathways. This function of HPSE is correlated to its expression level that is normally very low in majority of the tissues. Exceptionally, human platelets express high level of HPSE, suggesting a unique physiological role in this cell. Using K562 cell line, we found a progressive increase of HPSE during the megakaryocytic differentiation. Analysis of a series of megakaryocytic differentiation-related heparin-binding proteins (HBPs) in the cell culture medium revealed an exclusive positive correlation between the level of interleukin 6 (IL-6) and HPSE expression. IL-6 modulated megakaryocytic differentiation through activation of STAT3. Further, we demonstrated that overexpression of HPSE potentiates megakaryocytic differentiation, whereas elimination of HPSE led to a delayed differentiation. This function of HPSE is associated with its activity, as overexpression of inactive HPSE had no effect on IL-6 production and megakaryocytic differentiation. The role of HPSE is further supported by the observation in an umbilical cord blood CD34+ cells megakaryocytic differentiation model. Our data propose a novel role for HPSE in platelets production by a HPSE/IL-6/STAT3 positive feedback loop that specifically regulates megakaryocytes maturation.
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Lo, Wen-Cheng, Navneet Kumar Dubey, Feng-Chou Tsai, Jui-Hua Lu, Bou-Yue Peng, Pao-Chang Chiang, Abhinay Kumar Singh, Chia-Yu Wu, Hsin-Chung Cheng, and Win-Ping Deng. "Amelioration of Nicotine-Induced Osteoarthritis by Platelet-Derived Biomaterials Through Modulating IGF-1/AKT/IRS-1 Signaling Axis." Cell Transplantation 29 (January 1, 2020): 096368972094734. http://dx.doi.org/10.1177/0963689720947348.
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Besides inhalation, a few studies have indicated that the uptake of nicotine through air or clothing may be a significant pathway of its exposure among passive smokers. Nicotine is well known to exert various physiological impacts, including stimulating sympathetic nervous system, causing vascular disturbances, and inducing cell death. Therefore, we aimed to establish whether exposure of nicotine could induce articular cartilage degeneration in a mouse model of osteoarthritis (OA). We specifically assessed dose-dependent effect of nicotine in vitro to mimic its accumulation. Further, during the in vivo studies, mice subcutaneously administered with nicotine was examined for OA-associated pathologic changes. We found that nicotine significantly suppressed chondrocytes and chondrogenic markers (Sox, Col II, and aggrecan). Nicotine-treated mice also showed altered knee joint ultrastructure with reduced Col II and proteoglycans. After corroborating nicotine-induced OA characteristics, we treated this pathologic condition through employing platelet-derived biomaterial (PDB)-based regenerative therapy. The PDB significantly suppressed OA-like pathophysiological characteristics by 4 weeks. The mechanistic insight underlying this therapy demonstrated that PDB significantly restored levels of insulin-like growth factor 1 (IGF-1) signaling pathway proteins, especially pIGF-1 R, pAKT, and IRS-1, regulating extracellular matrix synthesis by chondrocytes. Taken together, the PDB exerts regenerative and reparative activities in nicotine-mediated initiation and progression of OA, through modulating IGF-1/AKT/IRS-1 signaling axis.
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Bauch, Juliane, and Andreas Faissner. "The Extracellular Matrix Proteins Tenascin-C and Tenascin-R Retard Oligodendrocyte Precursor Maturation and Myelin Regeneration in a Cuprizone-Induced Long-Term Demyelination Animal Model." Cells 11, no. 11 (May 28, 2022): 1773. http://dx.doi.org/10.3390/cells11111773.
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Oligodendrocytes are the myelinating cells of the central nervous system. The physiological importance of oligodendrocytes is highlighted by diseases such as multiple sclerosis, in which the myelin sheaths are degraded and the axonal signal transmission is compromised. In a healthy brain, spontaneous remyelination is rare, and newly formed myelin sheaths are thinner and shorter than the former ones. The myelination process requires the migration, proliferation, and differentiation of oligodendrocyte precursor cells (OPCs) and is influenced by proteins of the extracellular matrix (ECM), which consists of a network of glycoproteins and proteoglycans. In particular, the glycoprotein tenascin-C (Tnc) has an inhibitory effect on the differentiation of OPCs and the remyelination efficiency of oligodendrocytes. The structurally similar tenascin-R (Tnr) exerts an inhibitory influence on the formation of myelin membranes in vitro. When Tnc knockout oligodendrocytes were applied to an in vitro myelination assay using artificial fibers, a higher number of sheaths per single cell were obtained compared to the wild-type control. This effect was enhanced by adding brain-derived neurotrophic factor (BDNF) to the culture system. Tnr−/− oligodendrocytes behaved differently in that the number of formed sheaths per single cell was decreased, indicating that Tnr supports the differentiation of OPCs. In order to study the functions of tenascin proteins in vivo Tnc−/− and Tnr−/− mice were exposed to Cuprizone-induced demyelination for a period of 10 weeks. Both Tnc−/− and Tnr−/− mouse knockout lines displayed a significant increase in the regenerating myelin sheath thickness after Cuprizone treatment. Furthermore, in the absence of either tenascin, the number of OPCs was increased. These results suggest that the fine-tuning of myelin regeneration is regulated by the major tenascin proteins of the CNS.
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Karlsson, K., U. Lindahl, and S. L. Marklund. "Binding of human extracellular superoxide dismutase C to sulphated glycosaminoglycans." Biochemical Journal 256, no. 1 (November 15, 1988): 29–33. http://dx.doi.org/10.1042/bj2560029.
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The secretory enzyme extracellular superoxide dismutase (EC-SOD) occurs in at least three forms, which differ with regard to heparin affinity: A lacks affinity, B has intermediate affinity, and C has relatively strong affinity. The affinity of EC-SOD C for various sulphated glycosaminoglycans (GAGs) was assessed (a) by determining the concentration of NaCl required to release the enzyme from GAG-substituted Sepharose 4B and (b) by determining the relative potencies of the GAGs to release EC-SOD C from heparan sulphate-Sepharose 4B. Both methods indicated the same order of affinity. Heparin bound EC-SOD C about 10 times as avidly as the studied heparan sulphate preparation, which in turn was 10 and 150 times as efficient as dermatan sulphate and chondroitin sulphate respectively. Chondroitin sulphate showed weak interaction with EC-SOD C at physiological ionic strength. Heparin subfractions with high or low affinity for antithrombin III were equally efficient. The binding of EC-SOD C to heparin-Sepharose was essentially independent of pH in the range 6.5-9; below pH 6.5 the affinity increased, and beyond pH 9.5 there was a precipitous fall in affinity. The inhibitory effect of NaCl on the binding of EC-SOD C to GAGs indicates that the interaction is of electrostatic nature. EC-SOD C carries a negative net charge at neutral pH, and it is suggested that the binding occurs between the negative charges of the GAG sulphate groups and a structure in the C-terminal end of the enzyme that has a cluster of positive charges. These results are compatible with the notion that heparan sulphate proteoglycans on cell surfaces or in the intercellular matrix may serve to bind EC-SOD C in tissues.
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Feige, J. J., J. D. Bradley, K. Fryburg, J. Farris, L. C. Cousens, P. J. Barr, and A. Baird. "Differential effects of heparin, fibronectin, and laminin on the phosphorylation of basic fibroblast growth factor by protein kinase C and the catalytic subunit of protein kinase A." Journal of Cell Biology 109, no. 6 (December 1, 1989): 3105–14. http://dx.doi.org/10.1083/jcb.109.6.3105.
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Basic fibroblast growth factor (FGF) is synthesized as a phosphoprotein by both bovine capillary endothelial and human hepatoma cells in culture. Because basic FGF is characterized by its high affinity for heparin and its association in vivo with the extracellular matrix, we examined the possibility that the phosphorylation of this growth factor by purified protein kinase C (PK-C) and the catalytic subunit of cAMP-dependent protein kinase-A (PK-A) can be modulated by components of the extracellular matrix. Heparin and other glycosaminoglycans (GAGs) inhibit the ability of PK-C to phosphorylate basic FGF. In contrast, heparin can directly increase the phosphorylation of basic FGF by PK-A. While fibronectin, laminin, and collagen IV have no effect on the ability of PK-C to phosphorylate basic FGF, they all can inhibit the effects of PK-A. Thus, there is a differential effect of extracellular matrix-derived proteins and GAGs on the phosphorylation of basic FGF. The enhanced phosphorylation of basic FGF that is mediated by heparin is associated with a change in the kinetics of the reaction and the identity of the amino acid targeted by this enzyme. The amino acids that are targeted by PK-C and PK-A have been identified by phosphopeptide analyses as Ser64 and Thr112, respectively. In the presence of heparin, basic FGF is no longer phosphorylated by PK-A at the usual PK-A consensus site (Thr112), but instead is phosphorylated at the canonical PK-C site (Ser64). Accordingly, heparin inhibits the phosphorylation of basic FGF by PK-C presumably by masking the PK-C dependent consensus sequence surrounding Ser64. Thus, when basic FGF is no longer phosphorylated by PK-A in the receptor binding domain (Thr112), it loses the increased receptor binding ability that characterizes PK-A phosphorylated basic FGF. The results presented here demonstrate three novel features of basic FGF. First, they identify a functional effect of the binding of heparin to basic FGF. Second, they establish that the binding of heparin to basic FGF can induce structural changes that alter the substrate specificity of protein kinases. Third, and perhaps most important, the results demonstrate the existence of a novel interaction between basic FGF, fibronectin, and laminin. Although the physiological significance of this phosphorylation is not known, these results clearly suggest that the biological activities of basic FGF are regulated by a complex array of biochemical interactions with the proteins, proteoglycans, and glycosaminoglycans present in the extracellular milieu and the cytoplasm.
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Wadstein, Jan, Israel Sánchez Alvarez, and Lidia Bernal López. "Managing Skin Ageing as a Modifiable Disorder—The Clinical Application of Nourella® Dual Approach Comprising a Nano-Encapsulated Retinoid, Retilex-A® and a Skin Proteoglycan Replacement Therapy, Vercilex®." Cosmetics 9, no. 2 (March 15, 2022): 31. http://dx.doi.org/10.3390/cosmetics9020031.
Full textAbstract:
Skin ageing is a progressive, but modifiable, multi-factorial disorder that involves all the skin’s tissues. Due to its wide range of physiological and psychosocial complications, skin ageing requires rigorous clinical attention. In this review, we aim to encourage clinicians to consider skin ageing as a disorder and suggest a novel, dual approach to its clinical treatment. Topical retinoids and per-oral proteoglycans are promising, non-invasive, therapeutic modalities. To overcome the low bioavailability of conventional free retinoids, Nourella® cream with Retilex-A® (Pharma Medico, Aarhus, Denmark) was developed using a proprietary nano-encapsulation technology. The nano-encapsulation is a sophisticated ‘permeation/penetration enhancer’ that optimises topical drug delivery by increasing the surface availability and net absorption ratio. Treatment adherence is also improved by minimising skin irritation. Interventional evidence suggests the greater efficacy of Retilex-A® in improving skin thickness and elasticity compared with conventional free forms. It is also reported that the rejuvenating efficacy of Retilex-A® and tretinoin are comparable. Another skin anti-ageing approach is proteoglycan replacement therapy (PRT) with Vercilex®. Vercilex® in Nourella® tablet form has the potential to ameliorate proteoglycan dysmetabolism in aged skin by activating skin cells and improving collagen/elastin turnover. Replicated clinical trials evidenced that PRT can significantly enhance the density, elasticity and thickness of both intrinsically aged and photoaged skin. Evidently, Vercilex® and Retilex-A® share a range of bioactivities that underlie their synergistic activity, as observed in a clinical trial. Dual therapy with Nourella® tablets and cream produced greater effects on skin characteristics than monotherapy with each of the two treatments. In conclusion, Nourella® cream and tablets are safe and effective treatments for skin ageing; however, combining the two in a ‘dual skin rejuvenation system’ significantly improves treatment outcomes.
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McQuillan, D. J., C. J. Handley, M. A. Campbell, S. Bolis, V. E. Milway, and A. C. Herington. "Stimulation of proteoglycan biosynthesis by serum and insulin-like growth factor-I in cultured bovine articular cartilage." Biochemical Journal 240, no. 2 (December 1, 1986): 423–30. http://dx.doi.org/10.1042/bj2400423.
Full textAbstract:
The addition of foetal calf serum to explant cultures of adult bovine articular cartilage is known to stimulate proteoglycan synthesis in a dose-dependent manner. We have now shown the activity in serum responsible for this effect to be heat- and acid-stable, to be associated with a high-Mr complex in normal serum but converted to a low-Mr form under acid conditions. The activity has an apparent Mr approximately 10,000 and isoelectric points similar to those reported for insulin-like growth factors (IGFs). Addition of a monoclonal antibody against insulin-like growth factor-I (IGF-I) prevented foetal calf serum from stimulating proteoglycan synthesis. Physiological concentrations of recombinant IGF-I or pharmacological levels of insulin when added to cartilage cultures mimicked the proteoglycan-stimulatory activity of serum. IGF-I appeared to act by increasing the rate of proteoglycan synthesis and did not change the nature of the proteoglycan synthesized nor the rate of proteoglycan catabolism by the tissue, suggesting that IGF-I may be important in the regulation of proteoglycan metabolism in adult articular cartilage. Furthermore, IGF-I can replace foetal calf serum in the culture medium, thereby allowing the use of a fully-defined medium which will maintain the synthesis and tissue levels of proteoglycan in adult articular cartilage explants for up to 5 days.
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Kelkar, R., and G. A. Ateshian. "Contact Creep of Biphasic Cartilage Layers." Journal of Applied Mechanics 66, no. 1 (March 1, 1999): 137–45. http://dx.doi.org/10.1115/1.2789140.
Full textAbstract:
Integral transform methods are used to solve the contact creep problem between two identical cylindrical biphasic cartilage layers bonded to rigid impermeable subchondral bone substrates. The biphasic model employed for cartilage consists of a binary mixture of an incompressible porous-permeable solid phase and an incompressible fluid phase. Solutions are obtained as a function of time, from the instantaneous to the equilibrium responses of the tissue. A significant result of this analysis is that under application of a step load, fluid pressurization may support upward of 96 percent of the total applied load for more congruent joints, shielding the solid collagen-proteoglycan matrix of the tissue from excessive stresses during physiological loading durations. The protection imparted by interstitial fluid pressurization to the solid collagen-proteoglycan matrix of cartilage is investigated, and the influence of material properties and osteoarthritic changes on the potential loss of this protective effect is discussed.
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Stringer, S. E. "The role of heparan sulphate proteoglycans in angiogenesis." Biochemical Society Transactions 34, no. 3 (May 22, 2006): 451–53. http://dx.doi.org/10.1042/bst0340451.
Full textAbstract:
The presence of HS (heparan sulphate) proteoglycans on the cell surface and in the extracellular environment is critical to many physiological processes including the growth of new blood vessels from pre-existing vasculature (angiogenesis). A plethora of growth factors and their receptors, extracellular matrix molecules and enzymes bind to specific sites on the HS sugar chain. For example, HS proteoglycans have profound effects on the bioactivity of the key angiogenic factor VEGF (vascular endothelial growth factor) (VEGF165), affecting its diffusion, half-life and interaction with its tyrosine kinase receptors. A number of HS structural features that mediate the specific binding of VEGF165, including sulphation requirements, have been determined. In parallel, zebrafish embryos were used as a vertebrate model system to study the role in vascular development of the biosynthetic enzymes that create these specific binding sites on HS. It was discovered that knockdown of one of the HS 6-O-sulphotransferases in zebrafish with morpholino antisense oligonucleotides reduced vascular branching and corresponded to changes in the HS structure. The roles of the extracellular 6-O-sulphatase enzymes, the sulfs, in vascular development are now being investigated. Both oligosaccharides and small molecule biosynthetic enzyme inhibitors could be valuable HS-based strategies for controlling aberrant angiogenesis in diseases as diverse as cancer and heart disease.
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Fontanil, Tania, Yamina Mohamedi, Jorge Espina-Casado, Álvaro J. Obaya, Teresa Cobo, and Santiago Cal. "Hyalectanase Activities by the ADAMTS Metalloproteases." International Journal of Molecular Sciences 22, no. 6 (March 15, 2021): 2988. http://dx.doi.org/10.3390/ijms22062988.
Full textAbstract:
The hyalectan family is composed of the proteoglycans aggrecan, versican, brevican and neurocan. Hyalectans, also known as lecticans, are components of the extracellular matrix of different tissues and play essential roles in key biological processes including skeletal development, and they are related to the correct maintenance of the vascular and central nervous system. For instance, hyalectans participate in the organization of structures such as perineural nets and in the regulation of neurite outgrowth or brain recovery following a traumatic injury. The ADAMTS (A Disintegrin and Metalloprotease domains, with thrombospondin motifs) family consists of 19 secreted metalloproteases. These enzymes also perform important roles in the structural organization and function of the extracellular matrix through interactions with other matrix components or as a consequence of their catalytic activity. In this regard, some of their preferred substrates are the hyalectans. In fact, ADAMTSs cleave hyalectans not only as a mechanism for clearance or turnover of proteoglycans but also to generate bioactive fragments which display specific functions. In this article we review some of the physiological and pathological effects derived from cleavages of hyalectans mediated by ADAMTSs.
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Uhlin-Hansen, L., D. Langvoll, T. Wik, and SO Kolset. "Blood platelets stimulate the expression of chondroitin sulfate proteoglycan in human monocytes." Blood 80, no. 4 (August 15, 1992): 1058–65. http://dx.doi.org/10.1182/blood.v80.4.1058.1058.
Full textAbstract:
Abstract Mononuclear phagocytes synthesize chondroitin sulfate proteoglycan (CSPG), which is constitutively secreted. Because mononuclear phagocytes are known to interact with blood platelets, the effect of platelets on the release of CSPG in cultured human monocytes was investigated. After 6 days in vitro, the monocytes were supplied with fresh medium with different additions and subsequently exposed to [35S]sulfate for 24 hours before the medium fractions were harvested and analyzed for content of [35S]CSPG. Indirect evidence for the release of stimulatory factors from blood platelets was found when the addition of medium containing 50% serum made from platelet-rich plasma increased the expression of [35S]CSPG almost sevenfold compared with serum-free medium, whereas medium containing 50% serum made from platelet-depleted plasma increased the expression of [35S]CSPG about fourfold. Further, direct evidence for the stimulatory effect of platelets was found as the addition of autologous platelets to serum- free medium increased the expression of [35S]CSPG about threefold, and addition of supernatant from a corresponding number of thrombin- stimulated platelets was almost as efficient. The effect of five different platelet-derived factors (which are all present in serum) was investigated. Both platelet-derived growth factor (PDGF), platelet factor 4 (PF 4), and prostaglandin E2 (PGE2) used in physiologic concentrations were found to stimulate the expression of [35S]CSPG twofold to threefold, whereas transforming growth factor-beta had a slight inhibitory effect. 12-Hydroxyeicosatetraenoic acid had no significant effect on the expression of [35S]CSPG. Further evidence for the stimulatory effect of PDGF, PF 4, and PGE2 was found as serum depleted of these factors had significantly less stimulatory effect than control serum. The increased incorporation of [35S]sulfate into [35S]CSPG in cultures stimulated with serum or platelet-derived factors was not due to differences in molecular size or extent of sulfation of the proteoglycan molecules.
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Uhlin-Hansen, L., D. Langvoll, T. Wik, and SO Kolset. "Blood platelets stimulate the expression of chondroitin sulfate proteoglycan in human monocytes." Blood 80, no. 4 (August 15, 1992): 1058–65. http://dx.doi.org/10.1182/blood.v80.4.1058.bloodjournal8041058.
Full textAbstract:
Mononuclear phagocytes synthesize chondroitin sulfate proteoglycan (CSPG), which is constitutively secreted. Because mononuclear phagocytes are known to interact with blood platelets, the effect of platelets on the release of CSPG in cultured human monocytes was investigated. After 6 days in vitro, the monocytes were supplied with fresh medium with different additions and subsequently exposed to [35S]sulfate for 24 hours before the medium fractions were harvested and analyzed for content of [35S]CSPG. Indirect evidence for the release of stimulatory factors from blood platelets was found when the addition of medium containing 50% serum made from platelet-rich plasma increased the expression of [35S]CSPG almost sevenfold compared with serum-free medium, whereas medium containing 50% serum made from platelet-depleted plasma increased the expression of [35S]CSPG about fourfold. Further, direct evidence for the stimulatory effect of platelets was found as the addition of autologous platelets to serum- free medium increased the expression of [35S]CSPG about threefold, and addition of supernatant from a corresponding number of thrombin- stimulated platelets was almost as efficient. The effect of five different platelet-derived factors (which are all present in serum) was investigated. Both platelet-derived growth factor (PDGF), platelet factor 4 (PF 4), and prostaglandin E2 (PGE2) used in physiologic concentrations were found to stimulate the expression of [35S]CSPG twofold to threefold, whereas transforming growth factor-beta had a slight inhibitory effect. 12-Hydroxyeicosatetraenoic acid had no significant effect on the expression of [35S]CSPG. Further evidence for the stimulatory effect of PDGF, PF 4, and PGE2 was found as serum depleted of these factors had significantly less stimulatory effect than control serum. The increased incorporation of [35S]sulfate into [35S]CSPG in cultures stimulated with serum or platelet-derived factors was not due to differences in molecular size or extent of sulfation of the proteoglycan molecules.
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Vlodavsky, Israel, and Jin-ping Li. "Heparin, heparan sulfate and heparanase in inflammatory reactions." Thrombosis and Haemostasis 102, no. 11 (2009): 823–28. http://dx.doi.org/10.1160/th09-02-0091.
Full textAbstract:
SummaryHeparan sulfate (HS) proteoglycans at the cell surface and in the extracellular matrix of most animal tissues are essential in development and homeostasis, and are implicated in disease processes. Emerging evidence demonstrates the important roles of HS in inflammatory reactions, particularly in the regulation of leukocyte extravasation. Heparin, a classical anticoagulant, exhibits anti-inflammatory effects in animal models and in the clinic,presumably through interference with the functions of HS, as both polysaccharides share a high similarity in molecular structure. Apart of regulation during biosynthesis, the structures of HS and heparin are significantly modulated by heparanase, an endoglycosidase that is upregulated in a number of inflammatory conditions. Exploring the physiological roles of HS and heparin and the mode of heparanase action in modulating their functions during inflammation responses is of importance for future studies.
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Muhl, Lars, Etty Zwang, Neta Ilan, Yair Herishanu, Varda Deutsch, Elizabeth Naparstek, Israel Vlodavsky, Klaus Preissner, and Ben-Zion Katz. "Heparanase modulates heparinoids anticoagulant activities via non-enzymatic mechanisms." Thrombosis and Haemostasis 98, no. 12 (2007): 1193–99. http://dx.doi.org/10.1160/th07-04-0256.
Full textAbstract:
SummaryA key element for the physiological restriction of blood coagulation at the endothelial cell surface is its non-thrombogenic property, mainly attributed to cell surface heparan sulfate proteoglycans. Heparanase is an endo-β-D-glucuronidase with specific heparan sulfate degrading activity, which is produced and stored in platelets, and is released upon their activation. We examined the effects of heparanase pro-enzyme on coagulation functions, predominantly under physiological conditions. While heparanase pro-enzyme does not directly affect coagulation protein activities, it has profound effects on heparinoid-mediated regulation of coagulation responses, apparently via mechanisms that do not involve its enzymatic activity. Heparanase pro-enzyme reverses the anti-coagulant activity of unfractionated heparin on the coagulation pathway as well as on thrombin activity. In addition, heparanase pro-enzyme abrogated the factor X inhibitory activity of low-molecular-weight heparin (LMWH). The pro-coagulant effects of the non-active heparanase were also exerted by its major functional heparin-binding peptide. Finally, the effects of heparanase on the activity of factor VII activating protease that is auto-activated by heparinoids indicated a complete antagonistic action of heparanase in this system. Altogether, heparanase pro-coagulant activities that were also demonstrated in plasma samples from patients under LMWH treatment,point to a possible use of this molecule as antagonist for heparinoid treatment.
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Dunzendorfer, Stefan, Nicole Kaneider, Andrea Rabensteiner, Christian Meierhofer, Christina Reinisch, Jürgen Römisch, and Christian J. Wiedermann. "Cell-surface heparan sulfate proteoglycan–mediated regulation of human neutrophil migration by the serpin antithrombin III." Blood 97, no. 4 (February 15, 2001): 1079–85. http://dx.doi.org/10.1182/blood.v97.4.1079.
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Abstract The serpin antithrombin III (AT III) is reported to have hemostasis-regulating and anti-inflammatory properties. To determine its ability to influence thrombin-independent leukocyte responses, the direct effects of the AT III concentrate Kybernin P and a monoclonal antibody-purified AT III on neutrophil migration were studied. Chemotactic activity of human neutrophils isolated from the blood of healthy donors was determined in modified Boyden microchemotaxis chambers, and binding studies were performed according to standard experimental protocols. Preincubation in vitro of neutrophils with Kybernin P or immune-adsorbed AT III significantly deactivated migration toward fMet-Leu-Phe, or interleukin-8 (IL-8), in a concentration-dependent manner. In the absence of additional attractants, neutrophils exhibited a migratory response toward gradients of AT III preparations. True chemotaxis was confirmed in checkerboard assays. Analyses revealed that the AT III heparin-binding site interacts with neutrophil membrane–associated heparan sulfate proteoglycan receptors. Mechanisms of intracellular signaling differed; the deactivation of IL-8–induced chemotaxis resulted from tyrphostin-sensitive interactions of AT III-signaling with the IL-8 signal transduction pathway, whereas AT III–induced chemotaxis involved protein kinase C and phosphodiesterases. Signaling similarities between AT III and the proteoglycan syndecan-4 may suggest the binding of AT III to this novel type of membrane receptor. Under physiological conditions, AT III may prevent neutrophils from premature activation. Moreover, the systemic administration of AT III concentrate could have beneficial effects in combating systemic inflammation.
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ABRAHAMSSON, S.-O. "Exposure to Air during Surgery Inhibits Cellular Activity in Flexor Tendons." Journal of Hand Surgery 21, no. 3 (June 1996): 299–302. http://dx.doi.org/10.1016/s0266-7681(05)80188-3.
Full textAbstract:
In order to investigate the cellular effects of exposure to air during surgery and to compare the effects of simultaneous irrigation with physiological saline, the deep flexor tendons of both forepaws of 12 rabbits were surgically exposed. In one experiment, the extent of surgical exposure and, in a second experiment, the time of exposure was evaluated. Treated segments of the flexor tendons were collected and labelled in vitro for determination of the ability to synthesize DNA, proteoglycan, collagen and non-collagen protein. With increasing surgical exposure in vivo, an increasing rate of cellular proliferation was observed in segments of the exposed deep flexor tendons examined in vitro. Synthesis of matrix components and the rate of cellular proliferation were reduced by 50% after 40 to 100 minutes of exposure to air and by nearly 100% after 120 minutes of exposure. In contrast, irrigated tendons retained their cellular capacity to proliferate.
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Simsa-Maziel, Stav, Janna Zaretsky, Adi Reich, Yoav Koren, Ron Shahar, and Efrat Monsonego-Ornan. "IL-1RI participates in normal growth plate development and bone modeling." American Journal of Physiology-Endocrinology and Metabolism 305, no. 1 (July 1, 2013): E15—E21. http://dx.doi.org/10.1152/ajpendo.00335.2012.
Full textAbstract:
The proinflammatory cytokine interleukin-1 (IL-1) signals through IL-1 receptor type I (IL-1RI) and induces osteoclastogenesis and bone resorption mainly during pathological conditions. Little is known about the effect of excess or absence of IL-1 signaling on the physiological development of the growth plate and bone. In this study, we examine growth plate morphology, bone structure, and mechanical properties as well as osteoclast number in IL-1RI knockout mice to evaluate the role of IL-1RI in the normal development of the growth plate and bone. We show for the first time that IL-1RI knockout mice have narrower growth plates due to a smaller hypertrophic zone, suggesting a role for this cytokine in hypertrophic differentiation, together with higher proteoglycan content. The bones of theses mice exhibit higher trabecular and cortical mass, increased mineral density, and superior mechanical properties. In addition, IL-1RI knockout mice have significantly reduced osteoclast numbers in the chondro-osseous junction, trabecular bone, and cortical bone. These results suggest that IL-1RI is involved in normal growth plate development and ECM homeostasis and that it is significant in the physiological process of bone modeling.
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Lu, Y., K. H. Parker, and W. Wang. "Effects of osmotic pressure in the extracellular matrix on tissue deformation." Philosophical Transactions of the Royal Society A: Mathematical, Physical and Engineering Sciences 364, no. 1843 (April 18, 2006): 1407–22. http://dx.doi.org/10.1098/rsta.2006.1778.
Full textAbstract:
In soft tissues, large molecules such as proteoglycans trapped in the extracellular matrix (ECM) generate high levels of osmotic pressure to counter-balance external pressures. The semi-permeable matrix and fixed negative charges on these molecules serve to promote the swelling of tissues when there is an imbalance of molecular concentrations. Structural molecules, such as collagen fibres, form a network of stretch-resistant matrix, which prevents tissue from over-swelling and keeps tissue integrity. However, collagen makes little contribution to load bearing; the osmotic pressure in the ECM is the main contributor balancing external pressures. Although there have been a number of studies on tissue deformation, there is no rigorous analysis focusing on the contribution of the osmotic pressure in the ECM on the viscoelastic behaviour of soft tissues. Furthermore, most previous works were carried out based on the assumption of infinitesimal deformation, whereas tissue deformation is finite under physiological conditions. In the current study, a simplified mathematical model is proposed. Analytic solutions for solute distribution in the ECM and the free-moving boundary were derived by solving integro-differential equations under constant and dynamic loading conditions. Osmotic pressure in the ECM is found to contribute significantly to the viscoelastic characteristics of soft tissues during their deformation.
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Ethell, Iryna M., Kazuki Hagihara, Yoshiaki Miura, Fumitoshi Irie, and Yu Yamaguchi. "Synbindin, a Novel Syndecan-2–Binding Protein in Neuronal Dendritic Spines." Journal of Cell Biology 151, no. 1 (October 2, 2000): 53–68. http://dx.doi.org/10.1083/jcb.151.1.53.
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Dendritic spines are small protrusions on the surface of dendrites that receive the vast majority of excitatory synapses. We previously showed that the cell-surface heparan sulfate proteoglycan syndecan-2 induces spine formation upon transfection into hippocampal neurons. This effect requires the COOH-terminal EFYA sequence of syndecan-2, suggesting that cytoplasmic molecules interacting with this sequence play a critical role in spine morphogenesis. Here, we report a novel protein that binds to the EFYA motif of syndecan-2. This protein, named synbindin, is expressed by neurons in a pattern similar to that of syndecan-2, and colocalizes with syndecan-2 in the spines of cultured hippocampal neurons. In transfected hippocampal neurons, synbindin undergoes syndecan-2–dependent clustering. Synbindin is structurally related to yeast proteins known to be involved in vesicle transport. Immunoelectron microscopy localized synbindin on postsynaptic membranes and intracellular vesicles within dendrites, suggesting a role in postsynaptic membrane trafficking. Synbindin coimmunoprecipitates with syndecan-2 from synaptic membrane fractions. Our results show that synbindin is a physiological syndecan-2 ligand on dendritic spines. We suggest that syndecan-2 induces spine formation by recruiting intracellular vesicles toward postsynaptic sites through the interaction with synbindin.
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Wright, Malcolm, Paresh Jobanputra, Charles Bavington, Donald M. Salter, and George Nuki. "Effects of Intermittent Pressure-Induced Strain on the Electrophysiology of Cultured Human Chondrocytes: Evidence for the Presence of Stretch-Activated Membrane Ion Channels." Clinical Science 90, no. 1 (January 1, 1996): 61–71. http://dx.doi.org/10.1042/cs0900061.
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1. Cyclical pressurization of cultured chondrocytes results in increases in cyclic AMP and in the rate of proteoglycan synthesis. Intermittent increases in hydrostatic pressure are also associated with hyperpolarization of chondrocyte cell membranes and activation of Ca2+-dependent K+-ion channels but the physiological basis for this response to mechanical stimulation is unclear. 2. Experiments have been undertaken to better define the types of ion channels involved and to explore the possibility that the hyperpolarization response associated with cyclical pressurization of chondrocytes follows activation of stretch-activated ion channels. 3. The mean membrane potential of chondrocytes in non-confluent monolayer cell culture rose from −15.3 ± 0.24 mV to −21.1 ± 0.28 mV (n = 60, P < 0.0001) after intermittent pressurization (0.33 Hz, 16 kPa, 20 min). 4. Strain gauge measurements showed that cyclical pressurization was associated with strain on the base of the culture plate. The amplitude of the hyperpolarization response was proportional to the microstrain to which cells were subjected. 5. Membrane hyperpolarization did not occur when chondrocytes were subjected to cyclical pressurization in rigid glass culture dishes or plastic dishes positioned in the pressurization chamber so as to avoid bending of the base of the culture dish. 6. Indirect evidence that the hyperpolarization response after intermittent pressure-induced strain was associated with stimulation of stretch-activated ion channels was obtained from experiments with gadolinium, amiloride and hexamethylene amiloride, each of which abolished hyperpolarization. 7. Experiments with apamin, charybdotoxin and iberiotoxin showed that the Ca2+-activated K+ channels involved in the hyperpolarization response are apamin-sensitive, charybdotoxin- and iberiotoxin-resistant, low-conductance channels. 8. Somatostatin and cadmium chloride, which block L-type calcium channels, abolished strain-induced chondrocyte hyperpolarization. EGTA, which chelates extracellular Ca2+, reduced the response to 48% of control values, and thapsigargin, which raises intracellular Ca2+ by inhibition of Ca2+—ATPase in endoplasmic reticulum, caused hyperpolarization independently with further hyperpolarization after pressure-induced strain. These data indicate that chondrocyte hyperpolarization was dependent on intracellular Ca2+ concentrations. 9. Further work is required to determine whether stretch-activated ion channels shown to be associated with chondrocyte hyperpolarization after cyclical pressure-induced strain are also involved in the signal transduction process that leads to increases in proteoglycan synthesis.
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Medica, Pietro, Cristina Cravana, Alida Maria Ferlazzo, and Esterina Fazio. "Age-related functional changes of total thyroid hormones and glycosaminoglycans in growing calves." April-2020 13, no. 4 (2020): 681–86. http://dx.doi.org/10.14202/vetworld.2020.681-686.
Full textAbstract:
Background and Aim: During the physiological growing, thyroid and proteoglycan glycosaminoglycan (GAG) changes dynamically occur, according to genetic and non-genetic factors. The purpose of this research was to compare the effects of early postnatal development (10 days) until 210 days of life on the triiodothyronine (T3), thyroxine (T4), the relative T4:T3 ratio, and GAGs profile, and to define the different reference intervals of the calf's development through the various growing phases. Materials and Methods: The effect of growing on total thyroid hormones and GAG profiles was studied from 10 days to 210 days of age in 64 clinically healthy Brown calves, 30 males and 34 females. Blood samples were collected at 10, 20, 30, 60, 90, 120, 150, 180, and 210 days of age. Results: The results showed a significant effect of a calf's growth on T3, T4, and GAG values (p<0.0001). Significant correlations between T3 and T4 were observed. Compared to the previous time point, T3 showed a significant decrease at 20 days and at 60 days (p<0.01), while a significant increase was observed at 90 days and 210 days (p<0.05); T4 showed a significant decrease at 20 days (p<0.01), while significant increases were observed at both 180 days and 210 days (p<0.05); GAGs showed a significant increase at 120 days and 210 days (p<0.05). Positive and significant correlations between BW and GAGs in both males (p<0.0057) and females (p<0.0059) were observed. Conclusion: It can be concluded that the highest T3 and T4 concentrations have been associated with the early growing process (10 days), with an increasing trend also at 210 days, it is possible to hypothesize a probable metabolic effect of thyroid function in anabolic and/or catabolic directions during the calves' development. Likewise, it can be reasonably inferred that the highest plasma GAGs at 210 days may be due to their metabolic role during the development of growing calves. Taken together, these findings suggest the potential and relative contribution made by thyroid and GAGs effects on the dynamics of growing calves.
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Kunicki, Thomas J., Daniel Diaz, Shirley A. Williams, Richard W. Farndale, and Diane J. Nugent. "The Integrin α2 Dimorphism E534K Modulates Platelet Binding to Decorin but Not Collagen I,." Blood 118, no. 21 (November 18, 2011): 3256. http://dx.doi.org/10.1182/blood.v118.21.3256.3256.
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Abstract Abstract 3256 Background. Integrin a2b1-mediated adhesion to collagens supports cellular attachment, while decorin binding by a2b1 often attenuates adhesion. Collagen I binds to the a2 I-domain via the triple-helical sequence GFOGER, but the decorin binding site is not within the a2 I-domain and has not yet been identified. A single nucleotide polymorphism in the a2 gene ITGA2 (rs1801106) (G1600A) resulting in the amino acid substitution glutamate-534 to lysine-534 (E534K) is the basis for one of the most important human platelet alloantigen (HPA) systems, HPA-5, yet HPA-5 alleles do not influence the binding of a2b1 to collagen I, and the effect of HPA-5 on platelet function, if any, has not been determined. We sought to determine whether the minor allele HPA-5b (534K) might influence the a2b1-mediated adhesion of platelets to a physiologically relevant ligand other than collagen I. One such alternative ligand is decorin, an extracellular matrix small leucine (Leu)-rich proteoglycan (SLRP). Methods/Results. Two groups of normal subjects were compared. The first group (GG) consisted of ten donors homozygous for the major allele rs1801106G. The second group (GA+AA) included one donor homozygous for the minor allele rs1801106A and nine donors heterozygous for these alleles. Aside from this, there were no significant differences between the two groups with respect to platelet count, mean platelet volume, surface expression of four selected platelet receptors, GPIba, GPVI, aIIbb3 or a2b1, or the allelic distribution of five receptor genes, ITGA2B rs5911, ITGB3 rs5918, GP1BA rs6065, GP6 rs1613662 and P2RY1 rs1065776. In direct platelet adhesion assays, we determined that platelets from GG donors bound more strongly to decorin than those from GA+AA donors (p < 0.01), while platelets from either group bound equally well to collagen I (p = 0.73). Using platelets from donors homozygous for the major allele rs1801106G, adhesion to decorin was significantly inhibited by human alloantibodies specific for HPA-5a but not by GFOGER or an a2-specific monoclonal antibody 6F1 known to inhibit cell adhesion to collagen I. Conversely, GFOGER and 6F1 inhibited adhesion to collagen I but not decorin. Conclusions. The simplest explanation of our findings is that 534E, located in the a2 b-propeller domain, is necessary for maximal binding of a2b1 to decorin but not collagen I. These results suggest that 534E contributes to the decorin binding site and that this amino acid represents a potential target for interventions that can modify cell adhesion to decorin and perhaps other proteoglycans. This also represents the first instance of a functional difference attributable to the HPA-5 alleles. These results expand our understanding of the decorin and collagen I interactions described in murine and human studies of cell adhesion and cancer biology, where decorin and collagen I often have opposite effects that are both mediated by a2b1. Decorin knockout (Dcn(−/−)) murine embryonic fibroblasts exhibit greater adhesion to collagen and greater migration on collagen substrates, while decorin attenuates the aggressiveness and metastasis of tumor cells with diverse histologic backgrounds. Further studies of the relationship between the collagen I and decorin binding sites on integrin a2 are warranted, particularly regarding its influence on thrombosis and hemostasis. Disclosures: No relevant conflicts of interest to declare.
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Sottile, Jane, Feng Shi, Inna Rublyevska, Hou-Yu Chiang, Joseph Lust, and Jennifer Chandler. "Fibronectin-dependent collagen I deposition modulates the cell response to fibronectin." American Journal of Physiology-Cell Physiology 293, no. 6 (December 2007): C1934—C1946. http://dx.doi.org/10.1152/ajpcell.00130.2007.
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Communication between cells and the extracellular matrix (ECM) is critical for regulation of cell growth, survival, migration, and differentiation. Remodeling of the ECM can occur under normal physiological conditions, as a result of tissue injury, and in certain pathological conditions. ECM remodeling leads to alterations in ECM composition and organization that can alter many aspects of cell behavior, including cell migration. The cell migratory response varies depending on the type, amount, and organization of ECM molecules present, as well as the integrin and proteoglycan repertoire of the cells. We and others have shown that the deposition of several ECM molecules, including collagen types I and III, depends on the presence and stability of ECM fibronectin. Hence, the effect of fibronectin and fibronectin matrix on cell function may partially depend on its ability to direct the deposition of collagen in the ECM. In this study, we used collagen-binding fibronectin mutants and recombinant peptides that interfere with fibronectin-collagen binding to show that fibronectin-dependent collagen I deposition regulates the cell migratory response to fibronectin. These data show that the ability of fibronectin to organize other proteins in the ECM is an important aspect of fibronectin function and highlight the importance of understanding how interactions between ECM proteins influence cell behavior.
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Schönherr, Elke, Cord Sunderkötter, Renato V. Iozzo, and Liliana Schaefer. "Decorin, a Novel Player in the Insulin-like Growth Factor System." Journal of Biological Chemistry 280, no. 16 (February 8, 2005): 15767–72. http://dx.doi.org/10.1074/jbc.m500451200.
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Decorin is a multifunctional proteoglycan that is expressed by sprouting endothelial cells. Its expression supports capillary formation and cell survival. Previously, it was shown that some effects of decorin are mediated by protein kinase B and the cyclin-dependent kinase inhibitor, p21. However, the cell surface receptor responsible for these effects was unknown. We demonstrate that decorin binds to the insulin-like growth factor-I (IGF-I) receptor on endothelial cells with an affinity in the nanomolar range (KD= 18 nm), which is comparable with IGF-I (KD= 1.2 nm). Furthermore, decorin can bind IGF-I itself, but with a lower affinity (KD= 190 nm) than classical IGF-I-binding proteins. Decorin addition causes IGF-I receptor phosphorylation and activation, which is followed by receptor down-regulation. These effects are caused by the core protein of decorin, and the binding region could be mapped to the N terminus of the molecule. The physiological relevance of the decorin/IGF-I receptor interaction was corroborated in two animal models (e.g.inflammatory angiogenesis in the cornea and unilateral ureteral obstruction). In both models the IGF-I receptor was up-regulated in decorin-deficient mice compared with controls and the up-regulation could not compensate the decorin deficiency in the disease models. These data indicate that decorin is an important player in the IGF system and its loss cannot fully be compensated in different types of diseases.
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Takeno, Kenichi, Shigeru Kobayashi, Kohei Negoro, Kenzo Uchida, Tsuyoshi Miyazaki, Takafumi Yayama, Seiichiro Shimada, and Hisatoshi Baba. "Physical limitations to tissue engineering of intervertebral disc cells: effect of extracellular osmotic change on glycosaminoglycan production and cell metabolism." Journal of Neurosurgery: Spine 7, no. 6 (December 2007): 637–44. http://dx.doi.org/10.3171/spi-07/12/637.
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Object In this study, the authors examined how physiological levels of extracellular osmolality influence proteoglycan accumulation in nucleus pulposus cells in a 3D culture system. Methods Cells were isolated from the nucleus pulposus of caudal discs obtained from 18- to 24-month-old bovines. They were cultured for 6 days in alginate beads at 4 million cells/ml in Dulbecco modified Eagle medium containing 6% fetal bovine serum under 21% O2. Medium osmolality was altered by NaCl addition between 270 and 570 mOsm and monitored using a freezing point osmometer. The cell viability profile was determined by manual counting after trypan blue staining. Profiles across intact beads were determined by manual counting by using fluorescent probes and a transmission electron microscope. Lactate production was measured enzymatically, and glycosaminoglycan (GAG) accumulation was measured using a dimethylmethylene blue assay. Rate of sulfate GAG synthesis was measured using a standard [35S]sulfate radioactive method. Results The cell viability was similar for the high- and low-osmolality cultures. However, confocal microscopy showed that the cells were the largest at 270 mOsm and became smaller with increasing osmotic pressure. The GAG production was largest at 370 mOsm, the capacity for GAG production and cell metabolism (lactate production) was low under hypoosmolality and hyperosmolality, and cell death was observed on electron microscopy. Conclusions In the authors' model, the prevailing osmolality was a powerful regulator of GAG accumulation by cultured nucleus cells. Thus, these results indicate that GAG synthesis rates are regulated by GAG concentration, with implications both for the cause of degeneration and for tissue engineering.
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Salek-Ardakani, Shahram, John R. Arrand, David Shaw, and Mike Mackett. "Heparin and heparan sulfate bind interleukin-10 and modulate its activity." Blood 96, no. 5 (September 1, 2000): 1879–88. http://dx.doi.org/10.1182/blood.v96.5.1879.
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Abstract Glycosaminoglycans (GAG) are a group of negatively charged molecules that have been shown to bind and directly regulate the bioactivity of growth factors and cytokines such as basic fibroblast growth factor, transforming growth factor-β, IL-7, and interferon-γ. The ability of GAG to interact with human IL-10 (hIL-10) and the effect of these interactions on its biologic activity were analyzed. It was demonstrated by affinity chromatography that hIL-10 binds strongly to heparin–agarose at physiological pH. Biosensor-based binding kinetic analysis indicated an equilibrium dissociation constant, Kd, of 54 nmol/L for this interaction. Human IL-10 stimulated CD16 and CD64 expression on the monocyte/macrophage population within peripheral blood mononuclear cells, with optimal concentrations between 1 and 10 ng/mL. Soluble heparin, heparan sulfate, chondroitin sulfate, and dermatan sulfate were shown to inhibit the hIL-10–induced expression of CD16 and CD64 in a concentration-dependent manner. Heparin and heparan sulfate were most effective with IC50 values of 100 to 500 μg/mL. Considerably higher concentrations of dermatan sulfate and chondroitin 4-sulfate were required with an IC50 of 2000 to 5000 μg/mL, whereas chondroitin 6-sulfate was essentially inactive. The antagonistic effect of heparin on hIL-10 activity was shown to be dependent on N-sulfation, inasmuch as de-N-sulfated heparin had little or no inhibitory effect on the IL-10– induced expression of CD16, whereas the effect of de-O-sulfated heparin was comparable to that of unmodified heparin. Furthermore, the inhibition of cell-bound proteoglycan sulfation reduced the hIL-10–mediated expression of CD16 molecules on monocytes/macrophages. Taken together, these findings support the hypothesis that soluble and cell-surface GAG and, in particular, their sulfate groups are important in binding and modulation of hIL-10 activity.
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Salek-Ardakani, Shahram, John R. Arrand, David Shaw, and Mike Mackett. "Heparin and heparan sulfate bind interleukin-10 and modulate its activity." Blood 96, no. 5 (September 1, 2000): 1879–88. http://dx.doi.org/10.1182/blood.v96.5.1879.h8001879_1879_1888.
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Glycosaminoglycans (GAG) are a group of negatively charged molecules that have been shown to bind and directly regulate the bioactivity of growth factors and cytokines such as basic fibroblast growth factor, transforming growth factor-β, IL-7, and interferon-γ. The ability of GAG to interact with human IL-10 (hIL-10) and the effect of these interactions on its biologic activity were analyzed. It was demonstrated by affinity chromatography that hIL-10 binds strongly to heparin–agarose at physiological pH. Biosensor-based binding kinetic analysis indicated an equilibrium dissociation constant, Kd, of 54 nmol/L for this interaction. Human IL-10 stimulated CD16 and CD64 expression on the monocyte/macrophage population within peripheral blood mononuclear cells, with optimal concentrations between 1 and 10 ng/mL. Soluble heparin, heparan sulfate, chondroitin sulfate, and dermatan sulfate were shown to inhibit the hIL-10–induced expression of CD16 and CD64 in a concentration-dependent manner. Heparin and heparan sulfate were most effective with IC50 values of 100 to 500 μg/mL. Considerably higher concentrations of dermatan sulfate and chondroitin 4-sulfate were required with an IC50 of 2000 to 5000 μg/mL, whereas chondroitin 6-sulfate was essentially inactive. The antagonistic effect of heparin on hIL-10 activity was shown to be dependent on N-sulfation, inasmuch as de-N-sulfated heparin had little or no inhibitory effect on the IL-10– induced expression of CD16, whereas the effect of de-O-sulfated heparin was comparable to that of unmodified heparin. Furthermore, the inhibition of cell-bound proteoglycan sulfation reduced the hIL-10–mediated expression of CD16 molecules on monocytes/macrophages. Taken together, these findings support the hypothesis that soluble and cell-surface GAG and, in particular, their sulfate groups are important in binding and modulation of hIL-10 activity.
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Jahr, Holger, Anna E. van der Windt, Ufuk Tan Timur, Esther B. Baart, Wei-Shiung Lian, Bernd Rolauffs, Feng-Sheng Wang, and Thomas Pufe. "Physosmotic Induction of Chondrogenic Maturation Is TGF-β Dependent and Enhanced by Calcineurin Inhibitor FK506." International Journal of Molecular Sciences 23, no. 9 (May 4, 2022): 5110. http://dx.doi.org/10.3390/ijms23095110.
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Increasing extracellular osmolarity 100 mOsm/kg above plasma level to the physiological levels for cartilage induces chondrogenic marker expression and the differentiation of chondroprogenitor cells. The calcineurin inhibitor FK506 has been reported to modulate the hypertrophic differentiation of primary chondrocytes under such conditions, but the molecular mechanism has remained unclear. We aimed at clarifying its role. Chondrocyte cell lines and primary cells were cultured under plasma osmolarity and chondrocyte-specific in situ osmolarity (+100 mOsm, physosmolarity) was increased to compare the activation of nuclear factor of activated T-cells 5 (NFAT5). The effects of osmolarity and FK506 on calcineurin activity, cell proliferation, extracellular matrix quality, and BMP- and TGF-β signaling were analyzed using biochemical, gene, and protein expression, as well as reporter and bio-assays. NFAT5 translocation was similar in chondrocyte cell lines and primary cells. High supraphysiological osmolarity compromised cell proliferation, while physosmolarity or FK506 did not, but in combination increased proteoglycan and collagen expression in chondrocytes in vitro and in situ. The expression of the TGF-β-inducible protein TGFBI, as well as chondrogenic (SOX9, Col2) and terminal differentiation markers (e.g., Col10) were affected by osmolarity. Particularly, the expression of minor collagens (e.g., Col9, Col11) was affected. The inhibition of the FK506-binding protein suggests modulation at the TGF-β receptor level, rather than calcineurin-mediated signaling, as a cause. Physiological osmolarity promotes terminal chondrogenic differentiation of progenitor cells through the sensitization of the TGF-β superfamily signaling at the type I receptor. While hyperosmolarity alone facilitates TGF-β superfamily signaling, FK506 further enhances signaling by releasing the FKBP12 break from the type I receptor to improve collagenous marker expression. Our results help explain earlier findings and potentially benefit future cell-based cartilage repair strategies.
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Busiek, D. F., V. Baragi, L. C. Nehring, W. C. Parks, and H. G. Welgus. "Matrilysin expression by human mononuclear phagocytes and its regulation by cytokines and hormones." Journal of Immunology 154, no. 12 (June 15, 1995): 6484–91. http://dx.doi.org/10.4049/jimmunol.154.12.6484.
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Abstract Matrilysin is a recently described metalloproteinase with strong catalytic activity against a variety of extracellular matrix substrates including proteoglycans, elastin, laminin, fibronectin, gelatin, and entactin. Production of this metalloproteinase appears to be limited only to a few normal human cell types including glandular epithelium, mononuclear phagocytes, and renal mesangial cells. Furthermore, matrilysin expression in vivo has been demonstrated only in glandular epithelium, especially the endometrium. In the process of examining various cutaneous and lung inflammatory disorders for matrilysin expression by immunohistochemistry and in situ hybridization, we occasionally found monocytes within blood vessels and newly extravasated tissue-associated macrophages that exhibited matrilysin production. In specimens characterized by severe inflammation and, in particular, cystic fibrosis, this feature was commonly observed. We therefore studied the production of matrilysin by monocyte-derived macrophages in vitro in response to various physiologic signals such as endotoxin, phagocytosable material, cytokines, and hormones. We found that matrilysin expression was stimulated by LPS and opsonized zymosan. Up-regulation of matrilysin by LPS was PGE2-dependent, because indomethacin blocked production, an effect at least partially reversed by the addition of exogenous prostaglandin. LPS stimulated matrilysin production pretranslationally and, furthermore, when cultured cells were subjected to in situ hybridization after LPS exposure, considerable variability in matrilysin mRNA expression was observed on an individual cell basis, with some cells having strong signal and others being completely negative. We also found that matrilysin biosynthesis was inhibited by the lymphokines IL-4, IL-10, and IFN-gamma. Other cytokines such as IL-1, TNF-alpha, and IL-6 failed to modulate the production of matrilysin. Finally, matrilysin biosynthesis was suppressed by glucocorticoids and retinoids. Our studies indicate that matrilysin is produced in vivo by mononuclear phagocytes and is a highly regulated metalloproteinase whose production can be modified by a variety of physiologic and pharmacologic signals.
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Sun, D. D., X. E. Guo, M. Likhitpanichkul, W. M. Lai, and V. C. Mow. "The Influence of the Fixed Negative Charges on Mechanical and Electrical Behaviors of Articular Cartilage Under Unconfined Compression." Journal of Biomechanical Engineering 126, no. 1 (February 1, 2004): 6–16. http://dx.doi.org/10.1115/1.1644562.
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Unconfined compression test has been frequently used to study the mechanical behaviors of articular cartilage, both theoretically and experimentally. It has also been used in explant and gel-cell-complex studies in tissue engineering. In biphasic and poroelastic theories, the effect of charges fixed on the proteoglycan macromolecules in articular cartilage is embodied in the apparent compressive Young’s modulus and the apparent Poisson’s ratio of the tissue, and the fluid pressure is considered to be the portion above the osmotic pressure. In order to understand how proteoglycan fixed charges might affect the mechanical behaviors of articular cartilage, and in order to predict the osmotic pressure and electric fields inside the tissue in this experimental configuration, it is necessary to use a model that explicitly takes into account the charged nature of the tissue and the flow of ions within its porous interstices. In this paper, we used a finite element model based on the triphasic theory to study how fixed charges in the porous-permeable soft tissue can modulate its mechanical and electrochemical responses under a step displacement in unconfined compression. The results from finite element calculations showed that: 1) A charged tissue always supports a larger load than an uncharged tissue of the same intrinsic elastic moduli. 2) The apparent Young’s modulus (the ratio of the equilibrium axial stress to the axial strain) is always greater than the intrinsic Young’s modulus of an uncharged tissue. 3) The apparent Poisson’s ratio (the negative ratio of the lateral strain to the axial strain) is always larger than the intrinsic Poisson’s ratio of an uncharged tissue. 4) Load support derives from three sources: intrinsic matrix stiffness, hydraulic pressure and osmotic pressure. Under the unconfined compression, the Donnan osmotic pressure can constitute between 13%–22% of the total load support at equilibrium. 5) During the stress-relaxation process following the initial instant of loading, the diffusion potential (due to the gradient of the fixed charge density and the associated gradient of ion concentrations) and the streaming potential (due to fluid convection) compete against each other. Within the physiological range of material parameters, the polarity of the electric potential depends on both the mechanical properties and the fixed charge density (FCD) of the tissue. For softer tissues, the diffusion effects dominate the electromechanical response, while for stiffer tissues, the streaming potential dominates this response. 6) Fixed charges do not affect the instantaneous strain field relative to the initial equilibrium state. However, there is a sudden increase in the fluid pressure above the initial equilibrium osmotic pressure. These new findings are relevant and necessary for the understanding of cartilage mechanics, cartilage biosynthesis, electromechanical signal transduction by chondrocytes, and tissue engineering.
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Sweeney, Elizabeth A., Hugues Lortat-Jacob, Gregory V. Priestley, Betty Nakamoto, and Thalia Papayannopoulou. "Sulfated polysaccharides increase plasma levels of SDF-1 in monkeys and mice: involvement in mobilization of stem/progenitor cells." Blood 99, no. 1 (January 1, 2002): 44–51. http://dx.doi.org/10.1182/blood.v99.1.44.
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It was previously reported that treatment with the sulfated polysaccharide fucoidan or the structurally similar dextran sulfate increased circulating mature white blood cells and hematopoietic progenitor/stem cells (HPCs) in mice and nonhuman primates; however, the mechanism mediating these effects was unclear. It is reported here that plasma concentrations of the highly potent chemoattractant stromal-derived factor 1 (SDF-1) increase rapidly and dramatically after treatment with fucoidan in monkeys and in mice, coinciding with decreased levels in bone marrow. In vitro and in vivo data suggest that the SDF-1 increase is due to its competitive displacement from heparan sulfate proteoglycans that sequester the chemokine on endothelial cell surfaces or extracellular matrix in bone marrow and other tissues. Although moderately increased levels of interleukin-8, MCP1, or MMP9 were also present after fucoidan treatment, studies in gene-ablated mice (GCSFR−/−, MCP1−/−, or MMP9−/−) and the use of metalloprotease inhibitors do not support their involvement in the concurrent mobilization. Instead, SDF-1 increases, uniquely associated with sulfated glycan–mobilizing treatments and not with several other mobilizing agents tested, are likely responsible. To the authors' knowledge, this is the first published report of disrupting the SDF-1 gradient between bone marrow and peripheral blood through a physiologically relevant mechanism, resulting in mobilization with kinetics similar to other mobilizing CXC chemokines. The study further underscores the importance of the biological roles of carbohydrates.
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Suki, Béla, Satoru Ito, Dimitrije Stamenović, Kenneth R. Lutchen, and Edward P. Ingenito. "Biomechanics of the lung parenchyma: critical roles of collagen and mechanical forces." Journal of Applied Physiology 98, no. 5 (May 2005): 1892–99. http://dx.doi.org/10.1152/japplphysiol.01087.2004.
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The biomechanical properties of connective tissues play fundamental roles in how mechanical interactions of the body with its environment produce physical forces at the cellular level. It is now recognized that mechanical interactions between cells and the extracellular matrix (ECM) have major regulatory effects on cellular physiology and cell-cycle kinetics that can lead to the reorganization and remodeling of the ECM. The connective tissues are composed of cells and the ECM, which includes water and a variety of biological macromolecules. The macromolecules that are most important in determining the mechanical properties of these tissues are collagen, elastin, and proteoglycans. Among these macromolecules, the most abundant and perhaps most critical for structural integrity is collagen. In this review, we examine how mechanical forces affect the physiological functioning of the lung parenchyma, with special emphasis on the role of collagen. First, we overview the composition of the connective tissue of the lung and their complex structural organization. We then describe how mechanical properties of the parenchyma arise from its composition as well as from the architectural organization of the connective tissue. We argue that, because collagen is the most important load-bearing component of the parenchymal connective tissue, it is also critical in determining the homeostasis and cellular responses to injury. Finally, we overview the interactions between the parenchymal collagen network and cellular remodeling and speculate how mechanotransduction might contribute to disease propagation and the development of small- and large-scale heterogeneities with implications to impaired lung function in emphysema.
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Boyle, Kathleen A., and Teresa Compton. "Receptor-Binding Properties of a Soluble Form of Human Cytomegalovirus Glycoprotein B." Journal of Virology 72, no. 3 (March 1, 1998): 1826–33. http://dx.doi.org/10.1128/jvi.72.3.1826-1833.1998.
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ABSTRACT The human cytomegalovirus (HCMV) glycoprotein B (gB) (also known as gpUL55) homolog is an important mediator of virus entry and cell-to-cell dissemination of infection. To examine the potential ligand-binding properties of gB, a soluble form of gB (gB-S) was radiolabeled, purified, and tested in cell-binding experiments. Binding of gB-S to human fibroblast cells was found to occur in a dose-dependent, saturable, and specific manner. Scatchard analysis demonstrated a biphasic plot with the following estimated dissociation constants (Kd ): K d1, 4.96 × 10−6 M; K d2, 3.07 × 10−7 M. Cell surface heparan sulfate proteoglycans (HSPGs) were determined to serve as one class of receptors able to facilitate gB-S binding. Both HSPG-deficient Chinese hamster ovary (CHO) cells and fibroblast cells with enzymatically removed HSPGs had 40% reductions in gB-S binding, whereas removal of chondroitin sulfate had no effect. However, a significant proportion of gB-S was able to associate with the cell surface in the absence of HSPGs via an undefined nonheparin component. Binding affinity analysis of gB-S binding to wild-type CHO-K1 cells demonstrated biphasic binding kinetics (K d1, 9.85 × 10−6M; Kd2 , 4.03 × 10−8 M), whereas gB-S binding to HSPG-deficient CHO-677 cells exhibited single-component binding kinetics (Kd , 7.46 × 10−6 M). Together, these data suggest that gB-S associates with two classes of cellular receptors. The interaction of gB with its receptors is physiologically relevant, as evidenced by an inhibitory effect on HCMV entry when cells were pretreated with purified gB-S. This inhibition was determined to be manifested at the level of virus attachment. We conclude that gB is a ligand for HCMV that mediates an interaction with a cellular receptor(s) during HCMV infection.
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Fumagalli, Marta, Simona Daniele, Davide Lecca, Philip R. Lee, Chiara Parravicini, R. Douglas Fields, Patrizia Rosa, et al. "Phenotypic Changes, Signaling Pathway, and Functional Correlates of GPR17-expressing Neural Precursor Cells during Oligodendrocyte Differentiation." Journal of Biological Chemistry 286, no. 12 (January 4, 2011): 10593–604. http://dx.doi.org/10.1074/jbc.m110.162867.
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The developing and mature central nervous system contains neural precursor cells expressing the proteoglycan NG2. Some of these cells continuously differentiate to myelin-forming oligodendrocytes; knowledge of the destiny of NG2+ precursors would benefit from the characterization of new key functional players. In this respect, the G protein-coupled membrane receptor GPR17 has recently emerged as a new timer of oligodendrogliogenesis. Here, we used purified oligodendrocyte precursor cells (OPCs) to fully define the immunophenotype of the GPR17-expressing cells during OPC differentiation, unveil its native signaling pathway, and assess the functional consequences of GPR17 activation by its putative endogenous ligands, uracil nucleotides and cysteinyl leukotrienes (cysLTs). GPR17 presence was restricted to very early differentiation stages and completely segregated from that of mature myelin. Specifically, GPR17 decorated two subsets of slowly proliferating NG2+ OPCs: (i) morphologically immature cells expressing other early proteins like Olig2 and PDGF receptor-α, and (ii) ramified preoligodendrocytes already expressing more mature factors, like O4 and O1. Thus, GPR17 is a new marker of these transition stages. In OPCs, GPR17 activation by either uracil nucleotides or cysLTs resulted in potent inhibition of intracellular cAMP formation. This effect was counteracted by GPR17 antagonists and receptor silencing with siRNAs. Finally, uracil nucleotides promoted and GPR17 inhibition, by either antagonists or siRNAs, impaired the normal program of OPC differentiation. These data have implications for the in vivo behavior of NG2+ OPCs and point to uracil nucleotides and cysLTs as main extrinsic local regulators of these cells under physiological conditions and during myelin repair.
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Edwards, Danielle N., and Gregory J. Bix. "Roles of blood-brain barrier integrins and extracellular matrix in stroke." American Journal of Physiology-Cell Physiology 316, no. 2 (February 1, 2019): C252—C263. http://dx.doi.org/10.1152/ajpcell.00151.2018.
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Ischemicstroke is a leading cause of death and disability in the United States, but recent advances in treatments [i.e., endovascular thrombectomy and tissue plasminogen activator (t-PA)] that target the stroke-causing blood clot, while improving overall stroke mortality rates, have had much less of an impact on overall stroke morbidity. This may in part be attributed to the lack of therapeutics targeting reperfusion-induced injury after the blood clot has been removed, which, if left unchecked, can expand injury from its core into the surrounding at risk tissue (penumbra). This occurs in two phases of increased permeability of the blood-brain barrier, a physical barrier that under physiologic conditions regulates brain influx and efflux of substances and consists of tight junction forming endothelial cells (and transporter proteins), astrocytes, pericytes, extracellular matrix, and their integrin cellular receptors. During, embryonic development, maturity, and following stroke reperfusion, cerebral vasculature undergoes significant changes including changes in expression of integrins and degradation of surrounding extracellular matrix. Integrins, heterodimers with α and β subunits, and their extracellular matrix ligands, a collection of proteoglycans, glycoproteins, and collagens, have been modestly studied in the context of stroke compared with other diseases (e.g., cancer). In this review, we describe the effect that various integrins and extracellular matrix components have in embryonic brain development, and how this changes in both maturity and in the poststroke environment. Particular focus will be on how these changes in integrins and the extracellular matrix affect blood-brain barrier components and their potential as diagnostic and therapeutic targets for ischemic stroke.
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Sidis, Yisrael, Alan L. Schneyer, and Henry T. Keutmann. "Heparin and Activin-Binding Determinants in Follistatin and FSTL3." Endocrinology 146, no. 1 (January 1, 2005): 130–36. http://dx.doi.org/10.1210/en.2004-1041.
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Local regulation of pituitary FSH secretion and many other cellular processes by follistatin (FS) can be ascribed to its potent ability to bind and bioneutralize activin, in conjunction with binding to cell surface heparan-sulfate proteoglycans through a basic heparin-binding sequence (HBS; residues 75–86) in the first of the three FS domains. The FS homolog, FSTL3, also binds activin, but lacks any HBS and cannot associate with cell surfaces. We have used mutational analyses to define the determinants for heparin binding and activin interaction in FS and to determine the effects of conferring heparin binding to FSTL3. Mutants expressed from 283F cells were tested for cell surface and heparin affinity binding, for competititive activin binding and for bioactivity by suppression of pituitary cell FSH secretion. Replacement of the HBS or the full-length FS-domain 1 abolished cell surface binding but enhanced activin binding 4- to 8-fold. Surface binding was partially reduced after mutation of either lysine pair 75/76 or 81/82 and eliminated after mutation of both pairs. The 75/76 mutation reduced activin binding and, therefore, pituitary cell bioactivity by 5-fold. However, insertion of the HBS into FSTL3 did not restore heparin binding or pituitary-cell bioactivity. These results show that 1) the residues within the HBS are necessary but not sufficient for heparin binding, and 2) the HBS also harbors determinants for activin binding. Introduction of the full domain from FS conferred heparin binding to FSTL3, but activin binding was abolished. This implies an evolutionary safeguard against surface binding by FSTL3, supporting other evidence for physiological differences between FS and FSTL3.
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Bordin, S., B. Ghebrehiwet, and R. C. Page. "Participation of C1q and its receptor in adherence of human diploid fibroblast." Journal of Immunology 145, no. 8 (October 15, 1990): 2520–26. http://dx.doi.org/10.4049/jimmunol.145.8.2520.
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Abstract C1q binds through its collagen-like domain to specific surface receptors of fibroblasts and to adhesive elements of extracellular matrix including fibronectin, collagens, proteoglycans, and laminin. To determine whether C1q participates in fibroblast adhesion, cells in serum-free medium were plated on surfaces coated with purified C1q at physiologic ionic strength and pH. Surfaces coated with fibronectin or collagen type I served as positive controls, and those coated with BSA were negative controls. Substratum-adsorbed C1q promoted fibroblast adhesion to a maximum of 73% of available cells within 90 min at 37 degrees C. Adhesion was C1q concentration dependent, saturable, specific, and dependent on the collagen-like domain of the molecule. De novo protein synthesis plays a role in adhesion: pretreatment of fibroblasts with cycloheximide reduced adherence about 50% of controls. Addition of exogenous fibronectin, collagen type I, or C1q as soluble mediators did not affect adhesion of the cycloheximide-treated cells to C1q substrate. Adhesion could be accounted for primarily, although not completely, by the C1q receptors. Antibodies raised against the Raji cell C1q receptors (alpha C1qR Ab) specifically inhibited fibroblast adhesion to C1q substrates about 60% of controls. The binding of fibroblasts to C1q substrates could be inhibited about 24% of controls with the GRGDTP cell recognition peptide. GRGDTP and alpha C1q Ab had an additive effect on adhesion that was inhibited 77 to 80% of controls. We conclude from these data that aggregated rather than monomeric C1q may be the natural ligand of the fibroblast C1q receptor, and the biologic function of the receptor in cells of the connective tissue may be cell adhesion.
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Buschmann, M. D., Y. A. Gluzband, A. J. Grodzinsky, and E. B. Hunziker. "Mechanical compression modulates matrix biosynthesis in chondrocyte/agarose culture." Journal of Cell Science 108, no. 4 (April 1, 1995): 1497–508. http://dx.doi.org/10.1242/jcs.108.4.1497.
Full textAbstract:
This study focuses on the effect of static and dynamic mechanical compression on the biosynthetic activity of chondrocytes cultured within agarose gel. Chondrocyte/agarose disks (3 mm diameter) were placed between impermeable platens and subjected to uniaxial unconfined compression at various times in culture (2-43 days). [35S]sulfate and [3H]proline radiolabel incorporation were used as measures of proteoglycan and protein synthesis, respectively. Graded levels of static compression (up to 50%) produced little or no change in biosynthesis at very early times, but resulted in significant decreases in synthesis with increasing compression amplitude at later times in culture; the latter observation was qualitatively similar to that seen in intact cartilage explants. Dynamic compression of approximately 3% dynamic strain amplitude (approximately equal to 30 microns displacement amplitude) at 0.01-1.0 Hz, superimposed on a static offset compression, stimulated radiolabel incorporation by an amount that increased with time in culture prior to loading as more matrix was deposited around and near the cells. This stimulation was also similar to that observed in cartilage explants. The presence of greater matrix content at later times in culture also created differences in biosynthetic response at the center versus near the periphery of the 3 mm chondrocyte/agarose disks. The fact that chondrocyte response to static compression was significantly affected by the presence or absence of matrix, as were the physical properties of the disks, suggested that cell-matrix interactions (e.g. mechanical and/or receptor mediated) and extracellular physicochemical effects (increased [Na+], reduced pH) may be more important than matrix-independent cell deformation and transport limitations in determining the biosynthetic response to static compression. For dynamic compression, fluid flow, streaming potentials, and cell-matrix interactions appeared to be more significant as stimuli than the small increase in fluid pressure, altered molecular transport, and matrix-independent cell deformation. The qualitative similarity in the biosynthetic response to mechanical compression of chondrocytes cultured in agarose gel and chondrocytes in intact cartilage further indicates that gel culture preserves certain physiological features of chondrocyte behavior and can be used to investigate chondrocyte response to physical and chemical stimuli in a controlled manner.