A comparative analysis of the differential spatial and temporal distributions of the large (aggrecan, versican) and small (decorin, biglycan, fibromodulin) proteoglycans of the intervertebral disc

This study provides a comparative analysis of the temporal and spatial distribution of 5 intervertebral disc (IVD) proteoglycans (PGs) in sheep. The main PGs in the 2 and 10 y old sheep groups were polydisperse chondroitin sulphate and keratan sulphate substituted species. Their proportions did not differ markedly either with spinal level or disc zone. In contrast, the fetal discs contained 2 slow migrating (by composite agarose polyacrylamide gel electrophoresis, CAPAGE), relatively monodisperse chondroitin sulphate-rich aggrecan species which were also identified by monoclonal antibody 7-D-4 to an atypical chondroitin sulphate isomer presentation previously found in chick limb bud, and shark cartilage. The main small PG detectable in the fetal discs was biglycan, whereas decorin predominated in the 2 and 10 y old IVD samples; its levels were highest in the outer annulus fibrosus (AF). Versican was most abundant in the AF of the fetal sheep group; it was significantly less abundant in the 2 and 10 y old groups. Furthermore, versican was immunolocalised between adjacent layers of annular lamellae suggesting that it may have some role in the provision of the viscoelastic properties to this tissue. Versican was also diffusely distributed throughout the nucleus pulposus of fetal IVDs, and its levels were significantly lower in adult IVD specimens. This is the first study to identify versican in ovine IVD tissue sections and confirmed an earlier study which demonstrated that ovine IVD cells synthesised versican in culture (Melrose et al. 2000). The variable distribution of the PGs identified in this study provides further evidence of differences in phenotypic expression of IVD cell populations during growth and development and further demonstrates the complexity of the PGs in this heterogeneous but intricately organised connective tissue.     

Influence of strain on proteoglycan synthesis by valvular interstitial cells in three-dimensional culture

Differently loaded regions of the mitral valve contain distinct amounts and types of proteoglycans (PGs); these PG profiles are altered in abnormal loading and disease conditions. We developed an in vitro three-dimensional model to analyze PGs secreted by valvular interstitial cells (VICs) isolated from distinct regions of porcine mitral valves (leaflet or chordae) and subjected to either biaxial or uniaxial mechanical constraints. In addition, the PGs, DNA and collagen content of the collagen gels was monitored over time. All three PGs previously found in heart valves (decorin, biglycan and versican) were present in the collagen gels and the conditioned medium. Compared to unconstrained gels, the constrained collagen gels (whether biaxially or uniaxially loaded) retained more decorin and biglycan but less versican. However, the conditioned medium from constrained collagen gels contained higher amounts of all three PGs than did medium from unconstrained gels. Constrained collagen gels containing leaflet cells retained more decorin and biglycan than did those containing chordal cells. DNA content was maintained early in the culture period but was reduced by 55-80% after 7 days, whereas PG synthesis increased over time. At the end of the culture period, the cell density was highest in the biaxial region of gels seeded with leaflet cells. In contrast, collagen content in both constrained and unconstrained gels remained consistent over culture duration. This study provides valuable information about the role of applied loading on proteoglycan segregation, which should aid in tissue engineering applications and for understanding valve biology and pathology.     

Differential expression of proteoglycan epitopes and growth characteristics of intervertebral disc cells grown in alginate bead culture

This study assessed the growth characteristics and proteoglycan (PG) production in vitro of cells isolated from various zones; annulus fibrosus (AF), the transitional zone (TZ, an opalescent region situated between the AF and nucleus pulposus, NP) and the NP of the ovine intervertebral disc. The isolated cells were subsequently cultured in calcium alginate microspheres. Cellular metabolic activity was assessed over the 10 days of culture by the bioreduction of a tetrazolium dye substrate to a coloured formazan chromophore which could be measured colorimetrically at 490 nm. These data were correlated with the DNA content of beads measured using the fluorescent dye Hoechst 33258. Viable and non-viable cells in alginate beads were also assessed histochemically using the fluorescent dyes 5-chloromethylfluorescein diacetate and ethidium homodimer-1. PGs synthesised in culture were examined immunohistologically using monoclonal antibodies to defined glycosaminoglycan side chain PG epitopes. The results obtained clearly showed that AF and TZ cells differed from NP cells in the measured indices of cellular metabolism and in the extent of matrix deposition. In contrast to cells from the NP the cells from the AF and TZ were more metabolically active and programmed more to cellular proliferation than to matrix production.     

Identification and immunolocalization of chondroitin sulfate proteoglycans in tooth cementum

Proteoglycans (PGs) display a great diversity in their core proteins as well as carbohydrate structures and are thought to be involved in many biological functions. Recently we have identified and immunolocalized two keratan sulfate PGs, fibromodulin and lumican, in bovine tooth cementum (Cheng et al., Connect. Tissue Res. 34: 87-96, 1996). The objectives of this study were to identify and characterize chondroitin sulfate (CS) PGs in cementum. In order to explore their potential association with mineral, bovine cementum matrix molecules were fractionated into mineral-unbound and -bound matrices by sequential extraction. Both fractions were subjected to DEAE anion exchange column chromatography and the eluate collected was assayed for C4S and C6S isomers by dot blot immunoassay with specific monoclonal antibodies, 2-B-6 and 3-B-3, respectively. Two families of CSPGs were identified mainly in the mineral-unbound fraction. One contained only C4S glycosaminoglycan and the other both C6S and C4S. By biochemical and immunochemical analyses, decorin and biglycan were identified in the former and versican in the latter. The ratio of C6S to C4S isomers of cementum versican was approximately 7:1. Furthermore, these PGs were immunolocalized in and around tooth cementum using antibodies generated against the respective core proteins. Intensive immunostaining for versican was found almost exclusively in the lacunae housing cementocytes in cementum and osteocytes in alveolar bone, respectively. Immunostaining for decorin was mainly associated with collagen fibers in the periodontal ligament and slightly in cementum matrix, while the one for biglycan was mainly in cementoblasts/precementum. These differential tissue distributions of the CSPGs suggest that they may play distinct roles in cementogenesis.     

Proteoglycan distribution in lesions of atherosclerosis depends on lesion severity, structural characteristics, and the proximity of platelet-derived growth factor and transforming growth factor-beta.

The accumulation of proteoglycans (PGs) in atherosclerosis contributes to disease progression and stenosis and may partly depend on local regulation by growth factors such as platelet-derived growth factor (PDGF) and transforming growth factor (TGF)-beta. In this study, the distribution of the major extracellular PGs is compared with that of PDGF and TGF-beta isoforms in developing lesions of atherosclerosis from hypercholesterolemic nonhuman primates. Strong immunostaining for decorin, biglycan, versican, and hyaluronan is observed in both intermediate and advanced lesions. Perlecan staining is weak in intermediate lesions but strong in advanced lesions in areas bordering the plaque core. Immunostaining for PDGF-B and TGF-beta1 is particularly prominent in macrophages in intermediate and advanced lesions. In contrast, TGF-beta2 and TGF-beta3 and PDGF-A are present in both macrophages and smooth muscle cells. Overall, PG deposits parallel areas of intense growth factor immunostaining, with trends in relative localization that suggest interrelationships among certain PGs and growth factors. Notably, decorin and TGF-beta1 are distributed similarly, predominantly in the macrophage-rich core, whereas biglycan is prominent in the smooth muscle cell matrix adjoining TGF-beta1-positive macrophages. Versican and hyaluronan are enriched in the extracellular matrix adjacent to both PDGF- and TGF-beta1-positive cells. These data demonstrate that PG accumulation varies with lesion severity, structural characteristics, and the proximity of PDGF and TGF-beta.     

高压微胶囊成型装置制备用于成囊的海藻酸钙胶珠

利用高压微胶囊成型装置进行了制备用于成囊的海藻酸钙胶珠的试验研究 ,从而为用于药物控制释放的聚精氨酸基微胶囊的制备提供基础制备参数。将海藻酸钠溶液经试验装置滴入氯化钙溶液中固定化成海藻酸钙凝胶珠 ,考察了海藻酸钠浓度对制得的胶珠性能的影响及推进速度、针头内径、电压等对制得的胶珠直径的影响。得到较佳的海藻酸钠浓度为1.8% (W /V) ,发现推进速度提高 ,胶珠直径增大 :针头内径越大 ,胶珠直径越大 ;电压增大 ,胶珠的直径先减少后增大 ;不同针头内径制备出胶珠的平均直径最小时所对应的电压也随针头内径的增大而增大     

Immunohistochemical and histochemical characterization of the mucosubstances of odontogenic myxoma: histogenesis and differential diagnosis.

To discuss the dental origin of odontogenic myxoma and to provide further information for the differential diagnosis between this tumor and myxoid malignant fibrous histiocytoma (MFH) which occasionally occurs in jaw bones, the contents of glycosaminoglycans (GAGs) and proteoglycans (PGs) in the mucosubstances of 15 odontogenic myxomas, 5 myxoid MFH and 3 human fetal tooth germs in the bell stage of development were characterized using histochemical and immunohistochemical methods. Histochemical staining of hyaluronic acid (HA) was undertaken using biotinylated HA binding protein (B-HABP), and immunohistochemical detection was done using a panel of antibodies against chondroitin 6-sulfate (CS-6), chondroitin 4-sulfate (CS-4), dermatan sulfate (DS), keratan sulfate (KS), heparan sulfate (HS), aggrecan, PG-M/versican, decorin and biglycan. In odontogenic myxoma, CS-6, HA and PG-M/versican were observed in the myxomatous matrix of all cases, while KS and HS were seen in none. As for CS-4, DS, aggrecan, decorin and biglycan, only irregular and mild stainings were shown. Consistent and strong positive straining for CS-6, HA and PG-M/versican were seen in dental papilla and provided evidence supporting the origin of this tumor from dental papilla. Except for the constant staining for HA, the myxoid matrix was rarely stained for most GAGs and PGs in myxoid MFH. Immunodetection of CS-6 and PG-M/version with the use of monoclonal antibodies 3-B-3 and 2-B-1 is therefore recommended as a useful tool in differentiating odontogenic myoma from myxoid MFH.     

Mechanical strain enhances proteoglycan message in fibroblasts from asthmatic subjects

BACKGROUND: Remodelling of the asthmatic airway includes increased deposition of proteoglycan (PG) molecules. One of the stimuli driving airway remodelling may be excessive mechanical stimulation. OBJECTIVE: We hypothesized that fibroblasts from asthmatic patients would respond to excessive mechanical strain with up-regulation of message for PGs. METHODS: We obtained fibroblasts from asthmatic patients (AF) and normal volunteers (NF) using endobronchial biopsy. Cells were maintained in culture until the fifth passage and then grown on a flexible collagen-coated membrane. Using the Flexercell device, cells were then subjected to cyclic stretch at 30% amplitude at 1 Hz for 24 h. Control cells were unstrained. Total RNA was extracted from the cell layer and quantitative RT-PCR performed for decorin, lumican and versican mRNA. RESULTS: In unstrained cells, the expression of decorin mRNA was greater in AF than NF. With strain, NF showed increased expression of versican mRNA and AF showed increased expression of versican and decorin mRNA. The relative increase in versican mRNA expression with strain was greater in AF than NF. CONCLUSIONS: These data support the hypothesis that proteoglycan message is increased in asthmatic fibroblasts subject to mechanical strain. This finding has implications for the mechanisms governing airway wall remodelling in asthma.     

Pre-eclampsia-associated differential expression of proteoglycans in the umbilical cord arteries.

OBJECTIVE: The role of proteoglycans (PGs) of the umbilical cord arteries (UCAs) in the pathomechanism of pre-eclampsia is not known. Therefore we decided to compare the PGs of normal (control) UCAs and those of newborns delivered by mothers with pre-eclampsia. METHODS: PGs were extracted in dissociative conditions, purified by Q-Sepharose anion exchange chromatography and lyophilized. They were analyzed by gel filtration and SDS-PAGE before and after treatment with chondroitinase ABC. RESULTS: It was found that the PG preparation from pre-eclamptic UCAs had a higher amount of sulphated glycosaminoglycans (in relation to protein) than in the case of control UCAs. The predominant PG fraction included small PGs with core proteins of 45 and 47 kD, immunologically related to biglycan (45 kD) and decorin (45 and 47 kD). The expression of decorin core proteins was increased and that of biglycan slightly decreased in pre-eclamptic UCAs. Some other putative small PG core proteins (56, 53, 49, 42, 38 and 34 kD) were also found. They were present in higher amounts in pre-eclamptic UCAs. Larger PGs (core proteins of 99-110 and >150 kD), were detected in lower amounts, both in control and particularly in pre-eclamptic material. CONCLUSION: Pre-eclampsia is associated with alterations in PG composition of the UCAs. They may affect the mechanical properties of this organ and disturb fetal blood circulation.     

Three-dimensional growth of human hepatoma C3A cells within alginate beads for fluidized bioartificial liver

OBJECTIVES: Alginate beads allow cultivation of cells in a 3-dimensional environment. The aim of our study was to assess the influence of a 3-dimensional culture in alginate microbeads, on hepatic cell metabolism. METHODS: We used 2 types of alginate: low viscosity (LV) and medium viscosity (MV). The hepatic cell line C3A was encapsulated in alginate beads. Cells were cultured for 2 weeks. Using scanning electron microscopy, the morphology of 3D structures and the surfaces of cells were analyzed. Fluidized bed bioartificial liver experiments were performed 24 hours, 7, and 14 days after bead formation. RESULTS: Two different cell growth types in alginate beads were observed: channel-like structures and spherical aggregates characteristic of LV and MV alginate, respectively. A significant increase in albumin synthesis was observed in long-term culture. Formation of characteristic hepatic cell microvilli on cell surfaces was observed under scanning electron microscopy for both types of alginate. Prolonged static cultivation of C3A cells within the alginate beads in both types of alginates caused significant increases in albumin production in the fluidized bioreactor. CONCLUSIONS: Cultivation of the hepatic C3A cells within the alginate microbeads significantly improved bioreactor effectiveness in albumin production. The presence of extensions of cell membranes on the surface of hepatoma cells in 3-dimensional culture within the alginate beads indicated formation of microvilli-like structures characteristic of normal hepatocytes.     

Airway proteoglycans are differentially altered in fatal asthma

It has been suggested that airway remodelling is responsible for the persistent airway obstruction and decline in lung function observed in some asthmatic patients. The small airways are thought to contribute significantly to this functional impairment. Proteoglycans (PGs) are important components of the extracellular matrix (ECM) in the lungs. Besides controlling biophysical properties of the ECM, they play important roles in the regulation of some cytokines. Increased subepithelial PG deposition in the airways of mild asthmatics has been reported. However, there are no data on the PG content in small airways in asthma. This study has compared the content and distribution of PGs in large and small airways of patients who died of asthma with those in control lungs. Immunohistochemistry and image analysis were used to determine the content of lumican, decorin, biglycan, and versican in large (internal perimeter >6 mm) and small (internal perimeter < or =6 mm) airways of 18 patients who had died of asthma (A) and ten controls (C). The results were expressed as PG area (microm2)/epithelial basement membrane length (microm). The main differences between asthmatics and controls were observed in the small airways. There was a significant decrease in decorin and lumican contents in the external area of small airways in asthmatics (decorin: A = 1.05 +/- 0.27 microm, C = 3.97 +/- 1.17 microm, p = 0.042; lumican: A = 1.97 +/- 0.37 microm, C = 5.66 +/- 0.99 microm, p = 0.002). A significant increase in versican content in the internal area of small and large airways in asthmatics was also observed (small: A = 7.48 +/- 0.84 microm, C = 5.16 +/- 0.61 microm, p = 0.045; large: A = 18.38 +/- 1.94 microm, C = 11.90 +/- 2.86 microm, p = 0.028). The results show that PGs are differentially expressed in the airways of fatal asthma and may contribute to airway remodelling. These data reinforce the importance of the small airways in airway remodelling in asthma.     

Allergen-induced airway remodeling in brown norway rats: structural and metabolic changes in glycosaminoglycans

Increased proteoglycan (PG) deposition is a feature of airway remodeling in asthma. Glycosaminoglycans (GAGs) mediate many of the biological and mechanical properties of PGs by providing docking sites through their carbohydrate chains to bioactive ligands; therefore, it is imperative to define structural and metabolic changes of GAGs in asthma. Using a Brown Norway (BN) ovalbumin (OVA)-sensitized and -challenged rat model to induce airway remodeling, we found excessive deposition of chondroitin/dermatan (CS/DS)-, heparan (HS), and keratan (KS) sulfate GAGs in the airways and bronchoalveolar lavage cells of OVA-challenged rats. Disaccharide composition of CS/DS of OVA-challenged rats was significantly different compared with saline-treated (SAL) control rats, with increased levels of 0-, 6-, and 4-sulfated disaccharides. Increases in the amount and a change in the proportion of CS/DS versus HS GAGs were noted in OVA-challenged rats. The higher content and sulfation of CS/DS disaccharides was reflected by the increased expression of xylosyltransferase-I, β1,3-glucuronosyltransferase-I, chondroitin-4, and chondroitin-6 sulfotransferase genes and protein expression of xylosyltransferase-I and β1,3-glucuronosyltransferase-I in OVA-challenged rats. Genes encoding the core proteins of the CS/DS and KS-containing PGs, such as versican, biglycan, decorin, and lumican, were overexpressed in OVA-challenged rats. Our results suggest that GAG biosynthetic enzymes may be involved in the altered expression of GAGs in the airways and are potential targets for inhibiting excess PG-GAG deposition and the airway remodeling process in asthma.     

Effects induced by BMPS in cultures of human articular chondrocytes: Comparative studies

We compared anabolic and anti-catabolic activities of selected bone morphogenetic proteins (BMP-2, -4, -6, and -7) and cartilage-derived morphogenetic proteins (CDMP-1 and -2) in human normal adult articular chondrocytes. Ankle chondrocytes were cultured in alginate beads in the presence of 10% serum and treated with either growth factors only (each at 100 ng/ml) or the combination of interleukin-1 (IL-1β) (0.1 ng/ml) and BMPs. Chondrocyte metabolism was assessed by proteoglycan (PG) synthesis, content, DNA content, and cell survival. The results showed that BMP-2, -4, and -7 were more potent in stimulating PGs than other growth factors tested. The highest levels of PG synthesis were detected at day 9 in the presence of BMP-7. With regard to anti-catabolic properties, the effect depended upon treatment scheme (simultaneous or sequential). Under simultaneous cultures, BMP-2, -4, and -6 failed to counteract IL-1β induced inhibition of PG synthesis, while the CDMPs restored this parameter to serum control levels. Only BMP-7 showed consistent and pronounced anti-catabolic activity in either culture treatment scheme. None of the factors induced cell death or chondrocyte proliferation. In conclusion, the growth factors tested showed different levels of effects on human chondrocytes in culture, but only BMP-7 displayed both strong anabolic and anti-catabolic properties.     

Effects induced by BMPS in cultures of human articular chondrocytes: comparative studies

We compared anabolic and anti-catabolic activities of selected bone morphogenetic proteins (BMP-2, -4, -6, and -7) and cartilage-derived morphogenetic proteins (CDMP-1 and -2) in human normal adult articular chondrocytes. Ankle chondrocytes were cultured in alginate beads in the presence of 10% serum and treated with either growth factors only (each at 100 ng/ml) or the combination of interleukin-1 (IL-1 beta) (0.1 ng/ml) and BMPs. Chondrocyte metabolism was assessed by proteoglycan (PG) synthesis, content, DNA content, and cell survival. The results showed that BMP-2, -4, and -7 were more potent in stimulating PGs than other growth factors tested. The highest levels of PG synthesis were detected at day 9 in the presence of BMP-7. With regard to anti-catabolic properties, the effect depended upon treatment scheme (simultaneous or sequential). Under simultaneous cultures, BMP-2, -4, and -6 failed to counteract IL-1 beta induced inhibition of PG synthesis, while the CDMPs restored this parameter to serum control levels. Only BMP-7 showed consistent and pronounced anti-catabolic activity in either culture treatment scheme. None of the factors induced cell death or chondrocyte proliferation. In conclusion, the growth factors tested showed different levels of effects on human chondrocytes in culture, but only BMP-7 displayed both strong anabolic and anti-catabolic properties.     

In vivo maturation of scaffold-free engineered articular cartilage on hydroxyapatite

Tissue engineering is a promising approach, not only for cartilage, but also for osteochondral repair. Recent studies have demonstrated that scaffold-free cartilaginous tissue can be engineered using the alginate-recovered-chondrocyte (ARC) method. This method has also been applied to form osteochondral tissue using bovine articular chondrocytes and coralline hydroxyapatite (HA). The purpose of this study was to test whether osteochondral tissue, fabricated in vitro using the ARC method combined with a block of HA, would undergo maturation in vivo using a subcutaneous model in immunodeficient mice. Articular chondrocytes were isolated from the cartilage of New Zealand white rabbits and cultured in alginate beads. The cells with their associated matrix were recovered by dissolving the alginate beads with a sodium citrate buffer, resuspended in media and seeded onto a porous HA block. After 4 weeks of culture, some samples were analyzed, and others were implanted into subcutaneous pockets in nude mice. The analysis involved removing the cartilage portion of the de novo-formed ARC-HA graft and performing biochemical and histological examinations. Some samples were subjected to nondecalcified histology. Histological and immunohistochemical analyses of cartilaginous tissue were performed at 0, 2, 4, and 8 weeks after implantation. Biochemical characteristics were examined at 0, 4, and 8 weeks. The size and shape of the implanted ARC osteochondral tissue changed with time. The histological and immunohistochemical examination of the tissue revealed that it contained a cartilage-like matrix that stained strongly with Toluidine blue and for collagen type II. The proteoglycan (PG) content had increased significantly at 4 weeks from baseline. However, by 8 weeks, the PG content had decreased from 4 weeks. The results presented here represent a possible approach to form a tissue-engineered osteochondral implant. Further studies are needed to improve biomechanical properties of the osteochondral implant to be suitable for surgical transplantation.     

Effect of cyclic mechanical strain on glycosaminoglycan and proteoglycan synthesis by heart valve cells

Heart valves are presumed to remodel their extracellular matrix upon application of mechanical strains. In this study, we investigated the effect of cyclic tensile strain on valvular interstitial cells’ synthesis of glycosaminoglycans (GAGs) and proteoglycans (PGs), which are altered during myxomatous degeneration. Interstitial cells were isolated from mitral valve leaflets and chordate, and seeded separately within three-dimensional collagen gels. Cell-seeded collagen gels were then subjected to cyclic strains of 2%, 5% or 10% at 1.16 Hz for 48 h using a custom-built stretching device. The application of cyclic strains reduced the total GAGs retained within collagen gels in a magnitude-dependent manner for both leaflet and chordal cells. With increasing strain magnitude, however, secretion of total GAGs into the medium was reduced for leaflet cells and elevated for chordal cells. Retention of 4-sulfated GAGs increased with increasing strain magnitude for both cell types; for the chordal samples, retention of 6-sulfated GAGs was reduced at higher strain magnitudes. Compared to statically constrained or unconstrained conditions, the application of cyclic strain reduced the secretion of 6-sulfated GAGs by both cell types, and elevated secretion of 4-sulfated GAGs by leaflet cells only. Retention of the PG biglycan and secretion of the PG decorin was significantly reduced at 10% strain compared to 2% strain. In addition, there were numerous differences in the strain-dependent retention and secretion of GAGs and PGS within the leaflet and chordal groups. These results demonstrate that GAG and PG synthesis by VICs is regulated by cyclic stretching conditions.     

Covalent stabilization of alginate hydrogel beads via Staudinger ligation: assessment of poly(ethylene glycol) and alginate cross-linkers

Cellular encapsulation within alginate hydrogel capsules has broad applications in tissue engineering. In seeking to improve the inherent instability of ionically cross-linked alginate hydrogels, we previously demonstrated the covalent stabilization of Ba(2+) cross-linked alginate-azide beads via chemoselective Staudinger ligation using a 1-methyl-2-diphenylphosphino-terephthalate (MDT) terminated poly(ethylene glycol) (PEG) linker. In this study, we functionalized variant PEG, linear and branched, and alginate polymers with MDT groups to evaluate the effect of size, structural design, number of functional groups, and charge on the resulting hydrogel bead. All cross-linkers resulted in enhanced covalent stabilization of alginate beads, with significant decreases in swelling and resistance to dissolution via Ba(2+) chelation. The MDT-functionalized alginate resulted in the most stable and homogeneous bead, with the most restrictive permeability even after EDTA exposure. Co-encapsulation of MIN6 cells within the cross-linked alginate hydrogel beads resulted in minimal effects on viability, whereas the degree of proliferation following culture varied with cross-linker type. Altogether, the results illustrate that manipulating the cross-linker structural design permits flexibility in resulting alginate beads characteristics. Covalent stabilization of alginate hydrogel beads with these chemoselective alginate and PEG-based cross-linkers provides a unique platform for cellular encapsulation.     

Regional variation in human supraspinatus tendon proteoglycans: decorin, biglycan, and aggrecan

While tendons typically undergo primary tensile loading, the human supraspinatus tendon (SST) experiences substantial amounts of tension, compression, and shear in vivo. As a result, the functional roles of the extracellular matrix components, in particular the proteoglycans (PGs), are likely complex and important. The goal of this study was to determine the PG content in specific regions of the SST that exhibit differing mechanical function. The concentration of aggrecan, biglycan, and decorin was determined in six regions of the human SST using immunochemical techniques. We hypothesized that aggrecan concentrations would be highest in areas where the tendon likely experiences compression; biglycan levels would be highest in regions likely subjected to injury and/or active remodeling such as the anterior regions; decorin concentrations would be highest in regions of greatest tensile stiffness. Our results generally supported these hypotheses and demonstrated that aggrecan and biglycan share regional variability, with increased concentration in the anterior and posterior regions and smaller concentration in the medial regions. Decorin, however, was in high concentration throughout all regions. The data presented in this study represent the first regional measurements of PG in the SST. Together with our previous regional measurements of mechanical properties, these data can be used to evaluate SST structure-function relationships. With knowledge of the differences in specific PG content, their spatial variations in the SST, and their relationships to tendon mechanics, we can begin to associate defects in PG content with specific pathology, which may provide guidance for new therapeutic interventions.     

Reversible Secretion of Glycosaminoglycans and Proteoglycans by Cyclically Stretched Valvular Cells in 3D Culture

Mitral valve leaflets and chordae have been shown to contain different amounts and proportions of glycosaminoglycans (GAGs) and proteoglycans (PGs) corresponding to in vivo normal or diseased cyclic strain patterns. To understand the effect of cyclic strains on GAG/PG synthesis by valvular interstitial cells (VICs) isolated from valve leaflet and chordae separately, porcine VICs were seeded within collagen gels and alternately stretched or relaxed for 24 h periods for one week in a custom-designed tissue engineering bioreactor. We found cyclic-stretch-induced upregulation of total GAGs and of individual GAG classes secreted into the culture medium. Leaflet cells showed a delayed response to stretching compared to chordal cells, but altered the proportions of various GAG classes they secreted during the culture duration. Decorin and biglycan PGs were slightly responsive to stretch. We demonstrated that mechanical stretch and relaxation conditions reversibly regulate GAG and PG production in a novel 3D model of valve tissues. This is the first study using cyclic strains to modulate GAG/PG synthesis by valve cells and our results may have implications for the remodeling of the mitral valve as well as other tissues.