Relaxin and β-estradiol modulate targeted matrix degradation in specific synovial joint fibrocartilages: progesterone prevents matrix loss

Relaxin, a 6-kDa polypeptide hormone, is a potent mediator of matrix turnover and contributes to the loss of collagen and glycosaminoglycans (GAGs) from reproductive tissues, including the fibrocartilaginous pubic symphysis of several species. This effect is often potentiated by β-estradiol. We postulated that relaxin and β-estradiol might similarly contribute to the enhanced degradation of matrices in fibrocartilaginous tissues from synovial joints, which may help explain the preponderance of diseases of specific fibrocartilaginous joints in women of reproductive age. The objective of this study was to compare the in vivo effects of relaxin, β-estradiol, and progesterone alone or in various combinations on GAG and collagen content of the rabbit temporomandibular joint (TMJ) disc fibrocartilage, knee meniscus fibrocartilage, knee articular cartilage, and the pubic symphysis. Sham-operated or ovariectomized female rabbits were administered β-estradiol (20 ng/kg body weight), progesterone (5 mg/kg), or saline intramuscularly. This was repeated 2 days later and followed by subcutaneous implantation of osmotic pumps containing relaxin (23.3 μg/kg) or saline. Tissues were retrieved 4 days later and analyzed for GAG and collagen. Serum relaxin levels were assayed using enzyme-linked immunosorbent assay. Relaxin administration resulted in a 30-fold significant (p < 0.0001) increase in median levels (range, approximately 38 to 58 pg/ml) of systemic relaxin. β-estradiol, relaxin, or β-estradiol + relaxin caused a significant loss of GAGs and collagen from the pubic symphysis and TMJ disc and of collagen from articular cartilage but not from the knee meniscus. Progesterone prevented relaxin- or β-estradiol-mediated loss of these molecules. The loss of GAGs and collagen caused by β-estradiol, relaxin, or β-estradiol + relaxin varied between tissues and was most prominent in pubic symphysis and TMJ disc fibrocartilages. The findings suggest that this targeted modulation of matrix loss by hormones may contribute selectively to degeneration of specific synovial joints.     

Relaxin's induction of metalloproteinases is associated with the loss of collagen and glycosaminoglycans in synovial joint fibrocartilaginous explants

Diseases of specific fibrocartilaginous joints are especially common in women of reproductive age, suggesting that female hormones contribute to their etiopathogenesis. Previously, we showed that relaxin dose-dependently induces matrix metalloproteinase (MMP) expression in isolated joint fibrocartilaginous cells. Here we determined the effects of relaxin with or without beta-estradiol on the modulation of MMPs in joint fibrocartilaginous explants, and assessed the contribution of these proteinases to the loss of collagen and glycosaminoglycan (GAG) in this tissue. Fibrocartilaginous discs from temporomandibular joints of female rabbits were cultured in medium alone or in medium containing relaxin (0.1 ng/ml) or beta-estradiol (20 ng/ml) or relaxin plus beta-estradiol. Additional experiments were done in the presence of the MMP inhibitor GM6001 or its control analog. After 48 hours of culture, the medium was assayed for MMPs and the discs were analyzed for collagen and GAG concentrations. Relaxin and beta-estradiol plus relaxin induced the MMPs collagenase-1 and stromelysin-1 in fibrocartilaginous explants--a finding similar to that which we observed in pubic symphysis fibrocartilage, but not in articular cartilage explants. The induction of these proteinases by relaxin or beta-estradiol plus relaxin was accompanied by a loss of GAGs and collagen in joint fibrocartilage. None of the hormone treatments altered the synthesis of GAGs, suggesting that the loss of this matrix molecule probably resulted from increased matrix degradation. Indeed, fibrocartilaginous explants cultured in the presence of GM6001 showed an inhibition of relaxin-induced and beta-estradiol plus relaxin-induced collagenase and stromelysin activities to control baseline levels that were accompanied by the maintenance of collagen or GAG content at control levels. These findings show for the first time that relaxin has degradative effects on non-reproductive synovial joint fibrocartilaginous tissue and provide evidence for a link between relaxin, MMPs, and matrix degradation.     

Relaxin's induction of metalloproteinases is associated with the loss of collagen and glycosaminoglycans in synovial joint fibrocartilaginous explants

Diseases of specific fibrocartilaginous joints are especially common in women of reproductive age, suggesting that female hormones contribute to their etiopathogenesis. Previously, we showed that relaxin dose-dependently induces matrix metalloproteinase (MMP) expression in isolated joint fibrocartilaginous cells. Here we determined the effects of relaxin with or without β-estradiol on the modulation of MMPs in joint fibrocartilaginous explants, and assessed the contribution of these proteinases to the loss of collagen and glycosaminoglycan (GAG) in this tissue. Fibrocartilaginous discs from temporomandibular joints of female rabbits were cultured in medium alone or in medium containing relaxin (0.1 ng/ml) or β-estradiol (20 ng/ml) or relaxin plus β-estradiol. Additional experiments were done in the presence of the MMP inhibitor GM6001 or its control analog. After 48 hours of culture, the medium was assayed for MMPs and the discs were analyzed for collagen and GAG concentrations. Relaxin and β-estradiol plus relaxin induced the MMPs collagenase-1 and stromelysin-1 in fibrocartilaginous explants – a finding similar to that which we observed in pubic symphysis fibrocartilage, but not in articular cartilage explants. The induction of these proteinases by relaxin or β-estradiol plus relaxin was accompanied by a loss of GAGs and collagen in joint fibrocartilage. None of the hormone treatments altered the synthesis of GAGs, suggesting that the loss of this matrix molecule probably resulted from increased matrix degradation. Indeed, fibrocartilaginous explants cultured in the presence of GM6001 showed an inhibition of relaxin-induced and β-estradiol plus relaxin-induced collagenase and stromelysin activities to control baseline levels that were accompanied by the maintenance of collagen or GAG content at control levels. These findings show for the first time that relaxin has degradative effects on non-reproductive synovial joint fibrocartilaginous tissue and provide evidence for a link between relaxin, MMPs, and matrix degradation.     

The regional contribution of glycosaminoglycans to temporomandibular joint disc compressive properties

Understanding structure-function relationships in the temporomandibular joint (TMJ) disc is a critical first step toward creating functional tissue replacements for the large population of patients suffering from TMJ disc disorders. While many of these relationships have been identified for the collagenous fraction of the disc, this same understanding is lacking for the next most abundant extracellular matrix component, sulfated glycosaminoglycans (GAGs). Though GAGs are known to play a major role in maintaining compressive integrity in GAG-rich tissues such as articular cartilage, their role in fibrocartilaginous tissues in which GAGs are much less abundant is not clearly defined. Therefore, this study investigates the contribution of GAGs to the regional viscoelastic compressive properties of the temporomandibular joint (TMJ) disc. Chondroitinase ABC (C-ABC) was used to deplete GAGs in five different disc regions, and the time course for >95% GAG removal was defined. The compressive properties of GAG depleted regional specimens were then compared to non-treated controls using an unconfined compression stress-relaxation test. Additionally, treated and non-treated specimens were assayed biochemically and histologically to confirm GAG removal. Compared to untreated controls, the only regions affected by GAG removal in terms of biomechanical properties were in the intermediate zone, the most GAG-rich portion of the disc. Without GAGs, all intermediate zone regions showed decreased tissue viscosity, and the intermediate zone lateral region also showed a 12.5% decrease in modulus of relaxation. However, in the anterior and posterior band regions, no change in compressive properties was observed following GAG depletion, though these regions showed the highest compressive properties overall. Although GAGs are not the major extracellular matrix molecule of the TMJ disc, they are responsible for some of the viscoelastic compressive properties of the tissue. Furthermore, the mechanical role of sulfated GAGs in the disc varies regionally in the tissue, and GAG abundance does not always correlate with higher compressive properties. Overall, this study found that sulfated GAGs are important to TMJ disc mechanics in the intermediate zone, an important finding for establishing design characteristics for future tissue engineering efforts.     

Regional variation in the mechanical role of knee meniscus glycosaminoglycans

High compressive properties of cartilaginous tissues are commonly attributed to the sulfated glycosaminoglycan (GAG) fraction of the extracellular matrix (ECM), but this relationship has not been directly measured in the knee meniscus, which shows regional variation in GAG content. In this study, biopsies from each meniscus region (outer, middle, and inner) were either subjected to chondroitinase ABC (CABC) to remove all sulfated GAGs or not. Compressive testing revealed that GAG depletion in the inner and middle meniscus regions caused a significant decrease in modulus of relaxation (58% and 41% decreases, respectively, at 20% strain), and all regions exhibited a significant decrease in viscosity (outer: 29%; middle: 58%; inner: 62% decrease). Tensile properties following CABC treatment were unaffected for outer and middle meniscus specimens, but the inner meniscus displayed significant increases in Young's modulus (41% increase) and ultimate tensile stress (40% increase) following GAG depletion. These findings suggest that, in the outer meniscus, GAGs contribute to increasing tissue viscosity, whereas in the middle and inner meniscus, where GAGs are most abundant, these molecules also enhance the tissue's ability to withstand compressive loads. GAGs in the inner meniscus also contribute to reducing the circumferential tensile properties of the tissue, perhaps due to the pre-stress on the collagen network from increased hydration of the ECM. Understanding the mechanical role of GAGs in each region of the knee meniscus is important for understanding meniscus structure-function relationships and creating design criteria for functional meniscus tissue engineering efforts.     

Quantitative analysis and comparative regional investigation of the extracellular matrix of the porcine temporomandibular joint disc.

Characterization of the extracellular matrix of the temporomandibular joint (TMJ) disc is crucial to advancing efforts in tissue engineering the disc. However, the current literature is incomplete and often contradictory in its attempts to describe the nature of the TMJ disc matrix. The aim of this study was to identify the variation of key matrix components along the three axes of the porcine disc using ELISAs to quantify these matrix components, immunohistochemistry to identify their regional distribution, and SEM to examine collagen fiber diameter and orientation. The overall GAG content of the TMJ disc (including the dermatan sulfate proteoglycans) was 5.3+/-1.2% of the dry weight. Chondroitin sulfate, which comprised 74% of this total GAG content, was 4.4, 8.2, and 164 times more abundant than dermatan sulfate proteoglycan, keratan sulfate, and hyaluronic acid, respectively. In general, these GAGs were most concentrated in the intermediate zone of the TMJ disc, appearing in dense clusters, and least concentrated in the posterior band. Additionally, chondroitin sulfate was more abundant medially than laterally. Collagen II was discovered in trace amounts, with higher relative amounts in the intermediate zone. Collagen fibers were observed to run primarily in a ring-like fashion around the periphery of the disc and anteroposteriorly through the intermediate zone, with a mean fiber diameter of 18+/-9 mum. Characterization studies of the TMJ disc, including prior biomechanical and cell studies along with the current study of the extracellular matrix, collectively reveal a distinct character of the intermediate zone of the disc compared to its anterior and posterior bands.     

Effects of porcine relaxins upon uterine hypertrophy and protein metabolism in mice

Two variant forms of porcine relaxin (B and C) are active in producing relaxation of the guinea pig pubic symphysis and in effecting uterine growth in rats. Only relaxin B, however, is active in the mouse pubic ligament assay. These two hormones were compared in mice for their effects upon uterine growth and incorporation of radioactively labeled proline into soluble protein and collagen in vitro and in vivo. Both relaxin B and relaxin C produced an early (3-hr) elevation in in vitro protein synthesis and a later (6-hr) increase in collagen incorporation of proline at the time when the uterotrophic effect was maximal. In vivo effects of relaxin C on the uterus were in some cases greater than relaxin B in contrast to the complete inactivity of the former upon the pubic ligament of the mouse. These findings suggest a high degree of tissue specificity for relaxin stimulation, a variability in responsiveness among tissues in the same animal, and perhaps a primary role of relaxin in uterine function with pelvic relaxation representing a secondary function which has developed in certain species.     

Effects of the hormone relaxin on the metabolism of the glycosaminoglycans in the mouse symphysis pubis

The influence of the peptide hormone relaxin on the glycosaminoglycan (GAG) metabolism was investigated in the pubic ligament of the symphysis pubis and in serum of the virgin mouse. Fresh weight DNA and GAG content per 1 ligament is significantly increased, the level of water soluble protein is not affected. A shift in the electrophoretic GAG pattern by an increasing amount of hyaluronic acid and a decreasing amount of chondroitin sulfate and dermatan sulfate can be observed. Concerning GAG-splitting enzymes (N-acetylglucosaminidase, arylsulfatase, beta-glucuronidase) the N-acetylglucosaminidase reveals a significant increase of its activity in the interpubic ligament and in the serum. The data demonstrate that relaxin treatment induces some changes in the GAG metabolism.     

The distribution of different molecular species of collagen in fibrous, elastic and hyaline cartilages of the pig.

The distribution of type II collagen, considered to be characteristic of cartilaginous tissues, was determined in various specialized cartilages of the mature pig. The tissues examined were: (1) fibrocartilage of the semilunar meniscus of the knee; (2) elastic cartilage of the external ear; (3) hyaline cartilage of (a) the synovial joint (b) the thyroid plate of the larynx, and (c) the nasal septum. The predominant species of collagen in each tissue, whether type I or type II, was appraised semi-quantitatively by analysis of purified collagen solubilized by pepsin and of peptide fragments produced by cyanogen bromide. Cyanogen bromide-derived peptides were characterized by column chromatography on CM-cellulose and by electrophoresis in sodium dodecyl sulphate-polyacrylamide gels. The proportion of each type of collagen was determined precisely by isolating the homologous small peptides alpha1(II)CB6 [nomenclature of Miller (1973) Clin. Orthop. 92, 260-280], by column chromatography on phosphocellulose and determining their relative proportions by amino acid analysis. Thus collagen of the fibrocartilage of the meniscus proved to be all type I; type II was not detected. In contrast, collagen of elastic cartilage of the outer ear, after rigorous exclusion of perichondrium, was type II. Similarly, type II was the only collagen detected in all the mature hyalline cartilages examined.     

The low permeability of healthy meniscus and labrum limit articular cartilage consolidation and maintain fluid load support in the knee and hip

The knee meniscus and hip labrum appear to be important for joint health, but the mechanisms by which these structures perform their functions are not fully understood. The fluid phase of articular cartilage provides compressive stiffness and aids in maintaining a low friction articulation. Healthy fibrocartilage, the tissue of meniscus and labrum, has a lower fluid permeability than articular cartilage. In this study we hypothesized that an important function of the knee meniscus and the hip labrum is to augment fluid retention in the articular cartilage of a mechanically loaded joint. Axisymmetric hyperporoelastic finite element models were analyzed for an idealized knee and an idealized hip. The results indicate that the meniscus maintained fluid pressure and inhibited fluid exudation in knee articular cartilage. Similar, but smaller, effects were seen with the labrum in the hip. Increasing the fibrocartilage permeability relative to that of articular cartilage gave a consolidation rate and loss of fluid load support comparable to that predicted by meniscectomy or labrectomy. The reduced articular cartilage fluid pressure that was calculated for the joint periphery is consistent with patterns of endochondral ossification and osteophyte formation in knee and hip osteoarthritis. High articular central strains and loss of fluid load support after meniscectomy could lead to fibrillation. An intact low-permeability fibrocartilage is important for limiting fluid exudation from articular cartilage in the hip and knee. This may be an important aspect of the role of fibrocartilage in protecting these joints from osteoarthritis.     

Immunochemical and mechanical characterization of cartilage subtypes in rabbit.

Cartilage is categorized into three general subgroups, hyaline, elastic, and fibrocartilage, based primarily on morphologic criteria and secondarily on collagen (Types I and II) and elastin content. To more precisely define the different cartilage subtypes, rabbit cartilage isolated from joint, nose, auricle, epiglottis, and meniscus was characterized by immunohistochemical (IHC) localization of elastin and of collagen Types I, II, V, VI, and X, by biochemical analysis of total glycosaminoglycan (GAG) content, and by biomechanical indentation assay. Toluidine blue staining and safranin-O staining were used for morphological assessment of the cartilage subtypes. IHC staining of the cartilage samples showed a characteristic pattern of staining for the collagen antibodies that varied in both location and intensity. Auricular cartilage is discriminated from other subtypes by interterritorial elastin staining and no staining for Type VI collagen. Epiglottal cartilage is characterized by positive elastin staining and intense staining for Type VI collagen. The unique pattern for nasal cartilage is intense staining for Type V collagen and collagen X, whereas articular cartilage is negative for elastin (interterritorially) and only weakly positive for collagen Types V and VI. Meniscal cartilage shows the greatest intensity of staining for Type I collagen, weak staining for collagens V and VI, and no staining with antibody to collagen Type X. Matching cartilage samples were categorized by total GAG content, which showed increasing total GAG content from elastic cartilage (auricle, epiglottis) to fibrocartilage (meniscus) to hyaline cartilage (nose, knee joint). Analysis of aggregate modulus showed nasal and auricular cartilage to have the greatest stiffness, epiglottal and meniscal tissue the lowest, and articular cartilage intermediate. This study illustrates the differences and identifies unique characteristics of the different cartilage subtypes in rabbits. The results provide a baseline of data for generating and evaluating engineered repair cartilage tissue synthesized in vitro or for post-implantation analysis.     

Effects of co‐cultures of meniscus cells and articular chondrocytes on PLLA scaffolds

The knee meniscus, a fibrocartilaginous tissue located in the knee joint, is characterized by heterogeneity in extracellular matrix and biomechanical properties. To recreate these properties using a tissue engineering approach, co‐cultures of meniscus cells (MCs) and articular chondrocytes (ACs) were seeded in varying ratios (100:0, 75:25, 50:50, 25:75, and 0:100) on poly‐L ‐lactic acid (PLLA) scaffolds and cultured in serum‐free medium for 4 weeks. Histological, biochemical, and biomechanical tests were used to assess constructs at the end time point. Strong staining for collagen and glycosaminoglycan (GAG) was observed in all groups. Constructs with 100% MCs were positive for collagen I and constructs cultured with 100% ACs were positive for collagen II, while a mixture of collagen I and II was observed in other co‐culture groups. Total collagen and GAG per construct increased as the percentage of ACs increased (27 ± 8 µg, 0% AC to 45 ± 8 µg, 100% ACs for collagen and 12 ± 4 µg, 0% ACs to 40 ± 5 µg, 100% ACs for GAG). Compressive modulus (instantaneous and relaxation modulus) of the constructs was significantly higher in the 100% ACs group (63 ± 12 and 22 ± 9 kPa, respectively) when compared to groups with higher percentage of MCs. No differences in tensile properties were noted among groups. Specific co‐culture ratios were identified mimicking the GAG/DW of the inner (0:100, 25:75, and 50:50) and outer regions (100:0) of the meniscus. Overall, it was demonstrated that co‐culturing MCs and ACs on PLLA scaffolds results in functional tissue engineered meniscus constructs with a spectrum of biochemical and biomechanical properties. Biotechnol. Bioeng. 2009;103: 808–816. © 2009 Wiley Periodicals, Inc.     

Stimulation of ornithine decarboxylase by relaxin

Relaxin, a protein hormone of pregnancy, stimulated ornithine decarboxylase activity (EC 4.1.1.17) in two of its target tissues. Both the mouse public symphysis and uterus respond to a single injection of relaxin; within 2–4 hours after hormonal treatment of the mice, ornithine decarboxylase activity was observed to increase 2–8 fold over control levels. This increase in enzymatic activity may represent one step in the mechanism by which relaxin exerts its effects.     

The structure of the human symphysis pubis with special consideration of parturition and its sequelae

Review of the structure of the symphysis pubis, based on my extensive study of the pelvic joints ('31) shows changes from age, function, pregnancy hormones and stress of parturition. Primary physiologic shearing clefts and secondary traumatic clefts in cartilage are more frequent in females. Inter-digitations in the young osteocartilaginous border secure the vulnerable growth cartilage against increasing shearing forces. The retropubic eminence, ligamentous or cartilaginous, forms earlier in females, later, due to bony lipping in males, secondary to extrusion of disc cartilage. Ovarian and placental hormones in pregnancy cause remodeling and resorption of the posterior margin of the pubic facette and adjacent cortex, making a (variably) deep bony groove for greatly hypertrophied transverse ligaments. Delivery of a mature infant produces traumatic changes leading to extrusion of torn fibrocartilage in any direction, progressively loosening the symphysis, producing cartilage nodules, cysts and reactive bone formation. Older age degenerative arthritis is more frequent in parous females.     

Spatial distribution and orientation of dermatan sulfate in human medial collateral ligament

The proteoglycan decorin and its associated glycosaminoglycan (GAG), dermatan sulfate (DS), regulate collagen fibril formation, control fibril diameter, and have been suggested to contribute to the mechanical stability and material properties of connective tissues. The spatial distribution and orientation of DS within the tissue are relevant to these mechanical roles, but measurements of length and orientation from 2D transmission electron microscopy (TEM) are prone to errors from projection. The objectives of this study were to construct a 3D geometric model of DS GAGs and collagen fibrils, and to use the model to interpret TEM measurements of the spatial orientation and length of DS GAGs in the medial collateral ligament of the human knee. DS was distinguished from other sulfated GAGs by treating tissue with chondroitinase B, an enzyme that selectively degrades DS. An image processing pipeline was developed to analyze the TEM micrographs. The 3D model of collagen and GAGs quantified the projection error in the 2D TEM measurements. Model predictions of 3D GAG orientation were highly sensitive to the assumed GAG length distribution, with the baseline input distribution of 69+/-23 nm providing the best predictions of the angle measurements from TEM micrographs. The corresponding orientation distribution for DS GAGs was maximal at orientations orthogonal to the collagen fibrils, tapering to near zero with axial alignment. Sulfated GAGs that remained after chondroitinase B treatment were preferentially aligned along the collagen fibril. DS therefore appears more likely to bridge the interfibrillar gap than non-DS GAGs. In addition to providing quantitative data for DS GAG length and orientation in the human MCL, this study demonstrates how a 3D geometric model can be used to provide a priori information for interpretation of geometric measurements from 2D micrographs.     

The occurrence of hydroxyapatite crystals in extracellular matrix vesicles after surgical manipulation of the rabbit knee joint

Summary Free autologous grafts of synovial tissue were transplanted into experimental defects produced in the articular cartilage of rabbit knee joints. The grafted tissue underwent transformation into fibrocartilage. Extracellular matrix vesicles associated with calcified areas were present at the grafted sites. Hydroxyapatite crystals were found within these vesicles and in their vicinity. No calcification occurred in articular cartilage from sham operated joints in which defects were produced but no grafts made and in normal controls. These tissues showed abundant matrix vesicles devoid of crystalline mineral. A careful study of normal synovial tissue did not reveal matrix vesicles and calcifications. The present observations suggest that matrix vesicles in normal articular cartilage exist in a latent form. Vesicle mineralization following surgical manipulations of the joint is probably a manifestation of the metabolic stage of the tissue.     

Collagen studies in late pregnant relaxin null mice

The relaxin knockout (rlx -/-) mouse was used to assess the effect, during pregnancy, of relaxin with regard to water, collagen content, growth, and morphology of the nipple (N), vagina (V), uterus, cervix (C), pubic symphysis (PS), and mammary gland (MG). The results presented here indicate that during pregnancy, relaxin increases the growth of the N, C, V, and PS. Large increases in water content in the PS (20%) occurred in pregnant (Day 18.5) wild-type (rlx +/+) mice but not in rlx -/- animals. This indicates that in the PS, relaxin might increase the concentration of a water-retaining extracellular matrix component (hyaluronate). In the pregnant rlx +/+ mouse, collagen content decreased significantly in the N and V but not in other tissues. There were no significant changes in the rlx -/- mouse. This contrasts with findings in the rat, in which relaxin has been found to cause decreases in collagen concentrations in the V, C, and PS. Histological analysis showed that the collagen stain was more condensed in the tissues (V, C, PS, N, and MG) of rlx -/- mice than in those of rlx +/+ mice. This phenomenon indicates that the failure of collagen degradation and lack of growth in the N underlie the inability of the rlx -/- mice to feed their young, as reported previously. Vaginal and cervical luminal epithelia, which proliferated markedly in the rlx +/+ pregnant mice, remained relatively atrophic in the rlx -/- mice. As proliferation and differentiation of uterine and vaginal epithelia are thought to be induced by a paracrine stromal factor that acts upon estrogen stimulation, our results indicate that relaxin may be this paracrine factor.     

Growth factor effects on costal chondrocytes for tissue engineering fibrocartilage

Tissue-engineered fibrocartilage could become a feasible option for replacing tissues such as the knee meniscus or temporomandibular joint disc. This study employed five growth factors (insulin-like growth factor-I, transforming growth factor-beta1, epidermal growth factor, platelet-derived growth factor-BB, and basic fibroblast growth factor) in a scaffoldless approach with costal chondrocytes, attempting to improve biochemical and mechanical properties of engineered constructs. Samples were quantitatively assessed for total collagen, glycosaminoglycans, collagen type I, collagen type II, cells, compressive properties, and tensile properties at two time points. Most treated constructs had lower biomechanical and biochemical properties than the controls with no growth factors, suggesting a detrimental effect, but the treatment with insulin-like growth factor-I tended to improve the constructs. Additionally, the 6-week time point was consistently better than that at 3 weeks, with total collagen, glycosaminoglycans, and aggregate modulus doubling during this time. Further optimization of the time in culture and exogenous stimuli will be important in making a more functional replacement tissue.     

Lubricin Protects the Temporomandibular Joint Surfaces from Degeneration

The temporomandibular joint (TMJ) is a specialized synovial joint essential for the mobility and function of the mammalian jaw. The TMJ is composed of the mandibular condyle, the glenoid fossa of the temporal bone, and a fibrocartilagenous disc interposed between these bones. A fibrous capsule, lined on the luminal surface by the synovial membrane, links these bones and retains synovial fluid within the cavity. The major component of synovial fluid is lubricin, a glycoprotein encoded by the gene proteoglycan 4 (Prg4), which is synthesized by chondrocytes at the surface of the articular cartilage and by synovial lining cells. We previously showed that in the knee joint, Prg4 is crucial for maintenance of cartilage surfaces and for regulating proliferation of the intimal cells in the synovium. Consequently, the objective of this study was to determine the role of lubricin in the maintenance of the TMJ. We found that mice lacking lubricin have a normal TMJ at birth, but develop degeneration resembling TMJ osteoarthritis by 2 months, increasing in severity over time. Disease progression in Prg4 ^−/− mice results in synovial hyperplasia, deterioration of cartilage in the condyle, disc and fossa with an increase in chondrocyte number and their redistribution in clusters with loss of superficial zone chondrocytes. All articular surfaces of the joint had a prominent layer of protein deposition. Compared to the knee joint, the osteoarthritis-like phenotype was more severe and manifested earlier in the TMJ. Taken together, the lack of lubricin in the TMJ causes osteoarthritis-like degeneration that affects the articular cartilage as well as the integrity of multiple joint tissues. Our results provide the first molecular evidence of the role of lubricin in the TMJ and suggest that Prg4 ^−/− mice might provide a valuable new animal model for the study of the early events of TMJ osteoarthritis.     

Histochemistry for studying structure and function of the articular disc of the human temporomandibular joint

The articular disc of the temporomandibular joint (TMJ) is composed of fibrocartilage, and the extracellular matrix of this disc is composed mainly of collagen, glycosaminoglycan and proteoglycans. Research on the changes that occur in the composition of the articular disc of the TMJ is necessary for understanding the basis of the pathological process of internal derangement (ID), and a number of reports have been published in recent years on the application of refined histochemical techniques to investigate the structure and function of the TMJ. The direction of future TMJ disc studies should be towards obtaining more evidence to support previous results, and should hopefully be of practical use in terms of prevention and cure of ID.