Histochemical and immunohistochemical studies on the articular disk of the temporomandibular joint in rats

The effects of surgically induced malocclusion upon the articular disk of the temporomandibular joint in adult male rats were investigated histochemically and immunohistochemically using antibody against type II collagen. In the intermediate portion of the articular disk, the number of collagen fibers and chondrocytes was much less and the collagen fibers became more wavy, rough and irregular in surgically operated rats than in control rats. Clusters of chondrocytes were found in some experimental rats. The present experiments revealed that surgically induced malocclusion causes noticeable changes in collagen fibers and chondrocytes within the extracellular matrix of the articular disk of the temporomandibular joint.     

Age-related changes of elements in the human articular disk of the temporomandibular joint

To elucidate compositional changes of the articular disk (AD) of the temporomandibular joint (TMJ) by aging, elements of the ADs resected from 18 cadavers were determined by inductively coupled plasma atomic-emission spectrometry. It was found that calcium contents of ADs in TMJs increased progressively with aging, whereas the sulfur contents of the ADs decreased slightly with aging. Regarding the content of phosphorus, the contents increased progressively with aging. The study revealed that age-related changes of calcium contents in the ADs of TMJs were similar to those in women’s pubic symphyses, but not those in intervertebral disks and menisci.     

Anterior displacement of the TMJ disk: repositioning of the disk using a Mitek system. A 3D finite element study

In this paper the behaviors of the temporomandibular joint (TMJ) with an anteriorly displaced disk without reduction and with a surgically repositioned one were compared with the response of a healthy disk during jaw opening. The movement of each joint was obtained imposing the same opening path between incisors and assuming that the movement of the condyle is determined by the passive action of the masticatory muscles and the restrictions imposed by the articulating surfaces and the ligaments. A fiber-reinforced porohyperelastic model was used to simulate the behavior of the articular disk. The influence of the friction coefficient in the diseased joint was also analyzed, finding that the final displacement of the complex condyle-disk was smaller as the friction coefficient increased. On the other hand, its displacement in the repositioned joint was different than in the healthy case because the artificial sutures used in the surgery do not fully stabilize the disk posteriorly as the retrodiscal tissue does. The stress response of the disk changed in both pathologic cases: in the displaced joint the highest stresses moved from the intermediate zone (healthy case) to the posterior band, and in the reconstructed one the most loaded zone moved posteriorly at total opening. Besides, local stress concentrations appeared in the neighborhood of the artificial sutures and therefore damage of the disk and releasing of the sutures might be possible postoperatively.     

Cartilage interstitial fluid load support in unconfined compression

Under physiological conditions of loading, articular cartilage is subjected to both compressive strains, normal to the articular surface, and tensile strains, tangential to the articular surface. Previous studies have shown that articular cartilage exhibits a much higher modulus in tension than in compression, and theoretical analyses have suggested that this tension–compression nonlinearity enhances the magnitude of interstitial fluid pressurization during loading in unconfined compression, above a theoretical threshold of 33% of the average applied stress. The first hypothesis of this experimental study is that the peak fluid load support in unconfined compression is significantly greater than the 33% theoretical limit predicted for porous permeable tissues modeled with equal moduli in tension and compression. The second hypothesis is that the peak fluid load support is higher at the articular surface side of the tissue samples than near the deep zone, because the disparity between the tensile and compressive moduli is greater at the surface zone. Ten human cartilage samples from six patellofemoral joints, and 10 bovine cartilage specimens from three calf patellofemoral joints were tested in unconfined compression. The peak fluid load support was measured at 79±11% and 69±15% at the articular surface and deep zone of human cartilage, respectively, and at 94±4% and 71±8% at the articular surface and deep zone of bovine calf cartilage, respectively. Statistical analyses confirmed both hypotheses of this study. These experimental results suggest that the tension–compression nonlinearity of cartilage is an essential functional property of the tissue which makes interstitial fluid pressurization the dominant mechanism of load support in articular cartilage.     

Thrombospondin is present in articular cartilage and is synthesized by articular chondrocytes

Thrombospondin, a multifunctional adhesive glycoprotein originally identified in platelets, was isolated and identified from an extract of ovine articular cartilage. Immunoreactive material from a cartilage extract comigrated on gel electrophoresis with purified human platelet thrombospondin. When articular chondrocytes were cultured in the presence of 35S-methionine, metabolically labeled thrombospondin was immunoprecipitated from the culture medium and cell layer extract. These results demonstrate that thrombospondin is present in articular cartilage and is synthesized by articular chondrocytes.     

Mesenchymal progenitor cells in adult human articular cartilage

The transmembrane receptor Notch-1 regulates cell fate and differentiation and was suggested to identify a cell type with progenitor characteristics in newborn bovine articular cartilage. We show that Notch-1 is expressed on > 70% of BM-MSC in early passage monolayer culture. We also demonstrate that normal articular cartilage contains Notch-1+ cells and that the frequency is increased in OA. Most Notch-1+ cells in OA cartilage are located in the clusters of proliferating cells. These findings indicate that multipotential mesenchymal progenitor cells are present in articular cartilage from adult humans and that their frequency is increased in OA. This observation has implications for understanding the intrinsic repair capacity of articular cartilage and raises the possibility that these progenitor cells might be involved in the pathogenesis of arthritis.     

Three-dimensional orientations of talar articular surfaces in humans and great apes

The morphology of the talus prescribes relative positions and movements of the calcaneus and navicular with respect to the tibia, hence determining the overall geometry, mobility and function of the foot that mechanically interacts with environments. Clarifying the variations of the articular surface orientations of the talus in humans and extant great apes is therefore of importance in understanding the evolution of bipedal locomotion in the human lineage. The aim of this study is to clarify the three-dimensional orientations of three articular surfaces of the talus (superior, posterior calcaneal and navicular articular surfaces) by means of the newly proposed surface approximation method. Thirty-two tali in humans, chimpanzees, gorillas and orangutans were scanned using a three-dimensional noncontact digitizer, and the articular surfaces were then approximated using a paraboloid or a plane to calculate the orientations of the surfaces with respect to the body of the talus. The results quantitatively demonstrated that the superior articular surfaces in humans were relatively more parallel with the horizontal plane of the talar body, while those in apes were more medially oriented. Furthermore, the cylindrical axis defined by the shape of the posterior calcaneal articular surface was directed less anteroposteriorly in humans than in apes, in contrast to the fact that the subtalar axis is more anteroposteriorly oriented in humans. It was also demonstrated that the navicular articular surface in humans was more plantarly oriented and axially twisted. These specialized features of the human talus seem to be functionally linked to obligate bipedal locomotion. The talar morphological differences among the great apes were prominent in the mediolateral and rotational orientations of the navicular articular surfaces, possibly reflecting the degree of arboreality among the great apes.     

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.     

Production of sex steroid hormones from DHEA in articular chondrocyte of rats

Dehydroepiandrosterone (DHEA), a precursor of sex steroid hormones, is synthesized by cholesterol side-chain cleavage cytochrome P-450 and 17alpha-hydroxylase cytochrome P-450 mainly from cholesterol and converted to testosterone and estrogen by 3beta-hydroxysteroid dehydrogenase (3beta-HSD), 17beta-HSD, and aromatase cytochrome P-450. Although sex steroid hormones have important effects in the protection of articular cartilage, it is unclear whether articular cartilage has a local steroidogenic enzymatic machinery capable of metabolizing DHEA. This study was aimed to clarify whether steroidogenesis-related enzymes are expressed in articular chondrocytes, whether expression levels are changed by DHEA, and whether articular chondrocytes are capable of synthesizing sex steroid hormones from DHEA. Articular chondrocytes isolated from adult rats were cultured with DHEA for 3 days. All of the mRNA expressions of steroidogenesis-related enzymes were detected in cultured articular chondrocytes of rats, but the mRNA expression levels of testosterone and estradiol in cultured media increased after the addition of DHEA. These findings provided the first evidence that articular chondrocytes expressed steroidogenesis-related enzyme genes and that they are capable of locally synthesizing sex steroid hormones locally from DHEA.     

A novel terminal differentiation model of human articular chondrocytes in three-dimensional cultures mimicking chondrocytic changes in osteoarthritis

This study establishes a cell culture model mimicking the terminal differentiation occurring in osteoarthritic chondrocytes. Normal articular chondrocytes obtained from human knees treated with 5-azacytidine (Aza-C) were harvested 3, 7 and 14 days after treatment. Phenotypic and genetic changes of articular chondrocytes were detected. The results show that mRNA expression of collagen type II, a marker for normal functional articular chondrocytes, was significantly decreased after Aza-C treatment in comparison to the control cultures, while those of collagen type X and ALP, markers for hypertrophic chondrocytes, were significantly increased. Cell size and apoptotic rate of articular chondrocytes showed significant increases compared to the control after 14 days of Aza-C treatment. Terminal differentiation is shown by this model of three-dimensional cultured human articular chondrocytes, which could apply to the studies of the cellular mechanisms of osteoarthritis.     

Cell hypertrophy and type X collagen synthesis in cultured articular chondrocytes

Articular cartilage is a permanent tissue whose cells do not normally take part in the endochondral ossification process. To determine whether articular chondrocytes possess the potential to express traits associated with this process such as cell hypertrophy and type X collagen, chondrocytes were isolated from adult chicken tibial articular cartilage and maintained in long-term suspension cultures. As a positive control in these experiments, we used parallel cultures of chondrocytes from the caudal portion of chick embryo sternum. Both articular and sternal chondrocytes readily proliferated and progressively increased in size with time in culture. Many had undergone hypertrophy by 4-5 weeks. Analysis of medium-released collagenous proteins revealed that both articular and sternal chondrocytes initiated type X collagen synthesis between 3 and 4 weeks of culture; synthesis of this macromolecule increased with further growth. Immunofluorescence analysis of 5-week-old cultures showed that about 15% of articular chondrocytes and 30% of sternal chondrocytes produced type X collagen; strikingly, there appeared to be no obvious relationship between type X collagen production and cell size. The results of this study show that articular chondrocytes from adult chicken tibia possess the ability to express traits associated with endochondral ossification when exposed to a permissive environment. They suggest also that the process of cell hypertrophy and initiation of type X collagen synthesis are independently regulated both in articular and sternal chondrocytes.     

Lectin-binding in normal and osteoarthrotic articular cartilage from STR/1N-mouse knee joints

Fluorescein-isothiocyanate (FITC) labeled lectins were used to study the distribution pattern of specific binding-sites in histological sections of normal and osteoarthrotic articular cartilage from the mouse knee joint. Male inbred mice of the STR/1N-strain develop spontaneous arthrotic articular cartilage lesions on the medial condyle of tibia and femur. The varus-deformity of the knee joint leads to a recurrent medial patellar luxation with osteoarthrotic defects on the medial part of the facies patellaris femoris. It was demonstrated that the lectin staining pattern of osteoarthrotic articular cartilage, especially on the facies patellaris femoris, was different from that of normal articular cartilage. The differences in lectin staining corresponded to those observed between normal and fibrillated articular cartilage from human patellae. The normal articular cartilage of the mouse knee joint possessed lectin binding-sites for Concanavalin A (ConA) and wheat germ agglutinin (WGA), but not for Ulex europaeus agglutinin (UEA), soy bean agglutinin (SBA) and peanut agglutinin (PNA). In addition to the completely changed distribution pattern of ConA and WGA in osteoarthrotic cartilage, SBA, PNA and UEA developed distinct staining patterns particular to the fibrillated areas of arthrotic cartilage. The increased lectin-binding to arthrotic articular cartilage may be due to unmasking of sugars in the course of bondage breakdown in fibrillated cartilage or the production of pathological glycoproteins. It is evident that lectins can demonstrate minute differences between normal and arthrotic cartilage and it is concluded, therefore, that lectins are sensitive and specific tools for the study of degenerative joint diseases.     

The role of chondrocyte-matrix interactions in maintaining and repairing articular cartilage

Throughout life chondrocytes maintain the articular cartilage matrix by replacing degraded macromolecules and respond to focal cartilage injury or degeneration by increasing local synthesis activity. These observations suggest that mechanisms exist within articular cartilage that stimulate chondrocyte anabolic activity in response to matrix degradation or damage. An important cartilage anabolic factor, insulin-like growth factor I (IGF-I), appears to have a role in stimulating chondrocyte anabolic activity. Although IGF-I is ubiquitous, its bioavailability is controlled by a class of secreted proteins, IGF binding proteins (IGFPBs). Of the six known IGFPBs, IGFBP-3 is the most abundant in human articular cartilage. We recently found that with increasing age, articular chondrocytes increase their expression of IGFBP-3. This observation led us to investigate the potential role of IGFBP-3 in chondrocyte-matrix interactions. Using immunofluorescent staining and confocal microscopy we found that IGFBP-3 accumulates with increasing age in the chondrocyte territorial matrix where it co-localizes with fibronectin, but not with tenascin-C or type VI collagen. Using purified proteins we demonstrated that IGFBP-3 binds to fibronectin in a dose dependent manner, but not to tenascin-C. In vitro studies showed that IGFBP-3 alone inhibited chondrocyte synthetic activity while intact fibronectin alone significantly stimulated activity. When fibronectin and IGFBP-3 were combined we found that the inhibitory activity of low concentrations of IGFPB-3 was enhanced. These observations indicate that in mature articular cartilage IGF-I is stored in the chondrocyte territorial matrix through binding to a complex of IGFPB-3 and intact fibronectin. Storage of IGF-I of the territorial matrix may help maintain a relatively constant level of available IGF-I and the local increase in matrix synthesis following matrix damage may result from release of IGF-I. This mechanism may have an important role in maintaining and repairing articular cartilage and failure of this mechanism may lead to progressive articular cartilage degeneration.     

Variation in human limb joint articular morphology

Objectives

Synovial joints in human limbs strike a balance between mobility, stability, and articular fit, yet little is known about how these conflicting demands pattern intraspecific variation in articular shape. In this study, we use geometric morphometrics to establish the apportionment and magnitude of morphological variance of the articular surfaces of the human shoulder, elbow, hip, and knee. We hypothesize that variances will be comparable between articulating surfaces within a joint and will be larger in joints with smaller ranges of motion, given their plurality of functional demands.

Materials and Methods

Three-dimensional landmarks were taken on the articular surfaces of the glenohumeral, humeroulnar, acetabulofemoral, and tibiofemoral joints from CT scans of 200 skeletons from the University of Tennessee Donated Skeletal Collection (84 females, 116 males). Root mean-squared distances between articulations calculated from Procrustes shape coordinates were used to determine variance distributions.

Results

We found no difference in variances for each articular surface between the sexes or between left and right articular surfaces. A high range of motion is associated with greater morphological variance; however, this pattern is largely driven by the concave articular surfaces of each joint, which consistently exhibit statistically greater variance than their convex counterparts.

Discussion

The striking pattern of differential variance between articulating morphologies points to potential disparities in development between them. Consistently higher variance in concave surfaces may relate to chondral modeling theory for the formation of joints. Establishing intraspecific morphological variance patterns is a first step in understanding coordinated evolution among articular features.     

Toll-like receptor adaptor signaling molecule MyD88 on intervertebral disk homeostasis: in vitro, ex vivo studies

MyD88 is an adapter protein that links toll-like receptors (TLRs) and Interleukin-1 receptors (IL-1Rs) with downstream signaling molecules. The MyD88 has been found to be an essential mediator in the development of osteoarthritis in articular cartilage. However, the role of the MyD88 pathway has yet to be elucidated in the intervertebral disk (IVD). Using in vitro techniques, we analyzed the effect of MyD88 pathway-specific inhibition on the potent inflammatory and catabolic mediator LPS and IL-1 in bovine and human nucleus pulposus (NP) cells by assessing matrix-degrading enzyme expression, including matrix metalloproteases (MMPs) and a disintegrin-like and metalloprotease with thrombospondin motifs (ADAMTS family). We also analyzed inhibition of MyD88 in the regulation of inducible nitric oxide synthase and TLR-2. Finally, we used an ex vivo organ culture model to assess the effects of MyD88 inhibitor (MyD88i) on catabolic factor-induced disk degeneration in mice lumbar disks. In bovine NP cells, MyD88i potently antagonizes LPS- or IL-1-mediated induction of cartilage-degrading enzyme production, including MMP-1, MMP-13, ADAMTS-4, and ADAMTS-5. MyD88i also attenuates the LPS- or IL-1-mediated induction of iNOS and TLR-2 gene expression. Our ex vivo findings reveal inhibition of MyD88 via counteraction of IL-1-mediated proteoglycan depletion. The findings from this study demonstrate the potent anti-inflammatory and anti-catabolic effects of inhibition of MyD88 pathway inhibition on IVD homeostasis, suggesting a potential therapeutic benefit of a MyD88i in degenerative disk disease in the future.     

Ectopic mineralization of articular cartilage in the bullfrog Rana catesbeiana and its possible involvement in bone closure

Mineralization of the articular cartilage is a pathological condition associated with age and certain joint diseases in humans and other mammals. In this work, we describe a physiological process of articular cartilage mineralization in bullfrogs. Articular cartilage of the proximal and distal ends of the femur and of the proximal end of the tibia-fibula was studied in animals of different ages. Mineralization of the articular cartilage was detected in animals at 1 month post-transformation. This mineralization, which appeared before the hypertrophic cartilage showed any calcium deposition, began at a restricted site in the lateral expansion of the cartilage and then progressed to other areas of the epiphyseal cartilage. Mineralized structures were identified by von Kossa's staining and by in vivo incorporation of calcein green. Element analysis showed that calcium crystals consisted of poorly crystalline hydroxyapatite. Mineralized matrix was initially spherical structures that generally coalesced after a certain size to occupy larger areas of the cartilage. Alkaline phosphatase activity was detected at the plasma membrane of nearby chondrocytes and in extracellular matrix. Apoptosis was detected by the TUNEL (TDT-mediated dUTP-biotin nick end-labeling) reaction in some articular chondrocytes from mineralized areas. The area occupied by calcium crystals increased significantly in older animals, especially in areas under compression. Ultrastructural analyses showed clusters of needle-like crystals in the extracellular matrix around the chondrocytes and large blocks of mineralized matrix. In 4-year-old animals, some lamellar bone (containing bone marrow) occurred in the same area as articular cartilage mineralization. These results show that the articular cartilage of R. catesbeiana undergoes precocious and progressive mineralization that is apparently stimulated by compressive forces. We suggest that this mineralization is involved in the closure of bone extremities, since mineralization appears to precede the formation of a rudimentary secondary center of ossification in older animals.