The in vitro effects of four nutraceuticals, catechin hydrate, gallic acid, α-tocopherol and ascorbic acid, on the ability of human osteoarthritic chondrocytes of two female obese groups to form articular cartilage (AC) tissues and to reduce inflammation were investigated. Group 1 represented thirteen females in the 50–69 years old range, an average weight of 100 kg and an average body mass index (BMI) of 34⋅06 kg/m2. Group 2 was constituted of three females in the 70–80 years old range, an average weight of 75 kg and an average BMI of 31⋅43 kg/m2. The efficacy of nutraceuticals was assessed in monolayer cultures using histological, colorimetric and mRNA gene expression analyses. AC engineered tissues of group 1 produced less total collagen and COL2A1 (38-fold), and higher COL10A1 (2⋅7-fold), MMP13 (50-fold) and NOS2 (15-fold) mRNA levels than those of group 2. In comparison, engineered tissues of group 1 had a significant decrease in NO levels from day 1 to day 21 (2⋅6-fold), as well as higher mRNA levels of FOXO1 (2-fold) and TNFAIP6 (16-fold) compared to group 2. Catechin hydrate decreased NO levels significantly in group 1 (1⋅5-fold) while increasing NO levels significantly in group 2 (3⋅8-fold). No differences from the negative control were observed in the presence of other nutraceuticals for either group. In conclusion, engineered tissues of the younger but heavier patients responded better to nutraceuticals than those from the older but leaner study participants. Finally, cells of group 2 formed better AC tissues with less inflammation and better extracellular matrix than cells of group 1.

Epidemiological data show that osteoarthritis (OA) is significantly associated with lower birth weight, and that OA may be a type of fetal-originated adult disease. The present study aimed to investigate the prenatal food-restriction (PFR) effect on the quality of articular cartilage in female offspring to explore the underlying mechanisms of fetal-originated OA. Maternal rats were fed a restricted diet from gestational day (GD) 11 to 20 to induce intra-uterine growth retardation. Female fetuses and female adult offspring fed a post-weaning high-fat diet were killed at GD20 and postnatal week 24, respectively. Serum and knee cartilage samples from fetuses and adult female offspring were collected and examined for cholesterol metabolism and histology. Fetal serum corticosterone and insulin-like growth factor-1 (IGF-1) in the PFR group were lower than those of the control, but the serum cholesterol level was not changed. The lower expression of IGF-1 in the PFR group lasted into adulthood. The expression of extracellular matrix (ECM) genes, including type II collagen, aggrecan and cholesterol efflux genes including liver X receptor, were significantly induced, but the ATP-binding-cassette transporter A1 was unchanged. PFR could induce a reduction in ECM synthesis and impaired cholesterol efflux in female offspring, and eventually led to poor quality of articular cartilage and OA.

2-Oxoglutaric acid (2-Ox), a precursor to hydroxyproline – the most abundant amino acid in bone collagen, exerts protective effects on bone development during different stages of organism development; however, little is known about the action of 2-Ox on cartilage. The aim of the present study was to elucidate the influence of dietary 2-Ox supplementation on the growth plate, articular cartilage and bone of growing rats. A total of twelve male Sprague–Dawley rats were used in the study. Half of the rats received 2-oxoglutarate at a dose of 0·75 g/kg body weight per d in their drinking-water. Body and organ weights were measured. Histomorphometric analyses of the cartilage and bone tissue of the femora and tibiae were conducted, as well as bone densitometry and peripheral quantitative computed tomography (pQCT). Rats receiving 2-Ox had an increased body mass (P< 0·001) and absolute liver weight (P= 0·031). Femoral length (P= 0·045) and bone mineral density (P= 0·014), overall thickness of growth plate (femur P= 0·036 and tibia P= 0·026) and the thickness of femoral articular cartilage (P< 0·001) were also increased. 2-Ox administration had no effect on the mechanical properties or on any of the measured pQCT parameters for both bones analysed. There were also no significant differences in histomorphometric parameters of tibial articular cartilage and autofluorescence of femoral and tibial growth plate cartilage. Dietary supplementation with 2-Ox to growing rats exerts its effects mainly on cartilage tissue, having only a slight influence on bone. The effect of 2-Ox administration was selective, depending on the particular bone and type of cartilage analysed.

The uppermost superficial surface layer of articular cartilage, the ‘lamina splendens’ which provides a very low friction lubrication surface in articular joints, was investigated using atomic force microscopy (AFM). Complementary specimens were also observed under SEM at −10 °C without dehydration or sputter ion coating. Fresh adult pig osteochondral specimens were prepared from the patellas of pig knee joints and digested with the enzymes, hyaluronidase, chondroitinase ABC and alkaline protease. Friction coefficients between a pyrex glass plate and the osteochondral specimens digested by enzymes as well as natural (undigested) specimens were measured, using a thrust collar apparatus. Normal saline, hyaluronic acid (HA) and a mixture of albumin, globulin, HA (AGH) were used as lubrication media. The surface irregularities usually observed in SEM studies were not apparent under AFM. The articular cartilage surface was resistant to hyaluronidase and also to chondroitinase ABC, but a fibrous structure was exhibited in alkaline protease enzymes-digested specimens. AFM analysis revealed that the thickness of the uppermost superficial surface layer of articular cartilage was between 800 nm and 2 μm in adult pig articular cartilage. The coefficient of friction (c.f.) was significantly higher in chondroitinase ABC and alkaline protease enzymes digested specimens. Generally, in normal saline lubrication medium, c.f. was higher in comparison to HA and AGH lubrication media. The role of the uppermost, superficial surface layer of articular cartilage in the lubrication mechanism of joints is discussed.

The friction coefficients measured in diarthrodial joints are small. Theories of joint lubrication attribute this efficiency to entrapment or movement of synovial fluid, yet anatomical models of the surface are based on studies of isolated fragments of cartilage, not functional joints. To investigate the functional interrelationship of joint surfaces and synovial fluid, the ultrastructure of loaded joints was examined. Twenty-four New Zealand white rabbit knee joints were loaded either statically or moved ex vivo using simulated muscle forces and then plunge-frozen under load. After fixation in the frozen/loaded state by freeze-substitution fixation, the medial joint compartments were embedded in epoxy resin while still articulated. Bone was trimmed away from the articular surfaces, permitting the cartilage to be sectioned for light and electron microscopy. These joint surfaces were then compared with controls which were not loaded, not moved or had been disarticulated prior to embedding. Articular surfaces of loaded joints were smooth at magnifications from ×35 to ×7500, whereas the tibial surfaces of nonloaded joints were irregular. Small pools of joint fluid were observed at the meniscal edge and beneath the anterior horn of the meniscus. At magnifications of ×40000, the joint surfaces were separated by a uniform 100 nm space containing fluid. An amorphous, electron dense articular surface lamina was present but, when loaded, was thicker and flatter than previously reported. No surface pits or bumps were visible in embedded, loaded joints. This is the first ultrastructural study of intact loaded joints. The findings suggest that fluid film lubrication is present in diarthrodial joints, but the fluid sequestration postulated in several models is not apparent.

The subchondral bone plate supports the articular cartilage in diarthrodial joints. It has a significant mechanical function in transmitting loads from the cartilage into the underlying cancellous bone and has been implicated in the destruction of cartilage in osteoarthritis (OA) and its sparing in osteoporosis (OP), but little is known of its composition, structure or material properties. This study investigated the microscopic appearance and mineral composition of the subchondral bone plate in femoral heads from patients with OA or OP to determine how these correspond to changes in composition and stiffness found in other studies. Freeze-fractured full-depth samples of the subchondral bone plate from the femoral heads of patients with osteoarthritis, osteoporosis or a matched control group were examined using back scattered and secondary emission scanning electron microscopy. Other samples were embedded and polished and examined using back-scattered electron microscopy and electron probe microanalysis. The appearances of the samples from the normal and osteoporotic patients were very similar, with the subchondral bone plate overlayed by a layer of calcified cartilage. Osteoporotic samples presented a more uniform fracture surface and the relative thicknesses of the layers appeared to be different. In contrast, the OA bone plate appeared to be porous and have a much more textured surface. There were occasional sites of microtrabecular bone formation between the trabeculae of the underlying cancellous bone, which were not seen in the other groups, and more numerous osteoclast resorption pits. The calcified cartilage layer was almost absent and the bone plate was apparently thickened. The appearance of the osteoarthritic subchondral bone plate was, therefore, considerably different from both the normal and the osteoporotic, strongly indicative of abnormal cellular activity.

Articular cartilage undergoes cycles of compressive loading during joint movement, leading to its cyclical deformation and recovery. This loading is essential for chondrocytes to perform their normal function of maintenance of the extracellular matrix. Various lines of evidence suggest the involvement of the cytoskeleton in load sensing and response. The purpose of the present study is to describe the 3-dimensional (3D) architecture of the cytoskeleton of chondrocytes within their extracellular matrix, and to examine cytoskeletal responses to experimentally varied mechanical conditions. Uniformly sized explants of articular cartilage were dissected from adult rat femoral heads. Some were immediately frozen, cryosectioned and labelled for filamentous actin using phalloidin, and for the focal contact component vinculin or for vimentin by indirect immunofluorescence. Sections were examined by confocal microscopy and 3D modelling. Actin occurred in all chondrocytes, appearing as bright foci at the cell surface linked to an irregular network beneath the surface. Cell surface foci colocalised with vinculin, suggesting the presence of focal contacts between the chondrocyte and its pericellular matrix. Vimentin label occurred mainly in cells of the deep zone. It had a complex intracellular distribution, with linked networks of fibres surrounding the nucleus and beneath the plasma membrane. When cartilage explants were placed into organ culture, where in the absence of further treatments cartilage imbibes fluid from the culture medium and swells, cytoskeletal changes were observed. After 1 h in culture the vimentin cytoskeleton was disassembled, leading to diffuse labelling of cells. After a further hour in culture filamentous vimentin label reappeared in deep zone chondrocytes, and then over the next 48 h became more widespread in cells of the explants. Actin distribution was unaffected by culture. Further experiments were performed to test the effects of load on the cytoskeleton. Explants were placed in culture and immediately subjected to static uniaxial radially unconfined compressive loads of 0.5, 1, 2 or 4 MPa for 1 h using a pneumatic loading device. Loads greater than 0.5 MPa maintained the vimentin organisation over the culture period. At 0.5 MPa, the chondrocytes within the explant behaved as in free-swelling culture. The rapid change in vimentin organisation probably relates to rapid swelling of the explants—under free-swelling conditions, these reached their maximum swollen size in just 15 min of culture. The chondrocytes' response to change in tissue dimensions, and thus to their relationship to their immediate environment, was to disassemble their vimentin networks. Loading probably counteracts the swelling pressure of the tissue. Overall, this work suggests that chondrocytes maintain their actin cytoskeleton and modify their vimentin cytoskeleton in response to changing mechanical conditions.

The objective of this study was to investigate the normal distribution of cartilage thickness in the major joints of the lower limb in elderly individuals. A 12.5 MHz ultrasound transducer was used to measure the cartilage thickness in the right and left hip, knee and ankle joint of 10 individuals aged between 62 and 99 y. Distribution patterns of cartilage thickness were derived by b-spline interpolation and the average distribution computed in each surface. The maximum cartilage thickness in the hip joint was 2.6 (±0.36) mm and the mean thickness 1.3 (±0.17) mm. The CV% (a measure of thickness inhomogeneity within the joint surface) was 32%. In the knee, the maximal and mean values were 3.8 (±0.46) mm and 1.9 mm (±0.24) mm, respectively (CV%=34%), and in the ankle 1.7 (±0.25) mm and 1.0 (±0.16) mm (CV%=32%). Systematic differences existed between both sides in the knee, the distal femur showing a significantly greater thickness on the right. While the mean and maximal thicknesses were systematically higher in the knee than in the hip, and in the hip higher than in the ankle (P<0.05), there were no systematic differences in the thickness inhomogeneity of the 3 joints. Only the malleolus showed a somewhat more uniform thickness than the other joint surfaces. The variablity between individuals was similar for all joints for mean thickness, but the interindividual variability of the maximal thickness values was highest in the knee and lowest in the ankle. Whereas the cartilage thickness distributions in the joints of the lower limb have been suggested to reflect the pressure distribution within the articular surface, the absolute thickness is proposed to be a function of dynamic loading (range of motion) during gait, rather than being a reflection of the static articular pressure.

The fibrillar architecture in the general matrix of softened cartilage has been compared with that of the normal matrix using both Nomarski light microscopy and transmission electron microscopy with combined stereoscopic reconstruction. A pseudorandom network developed from an overall radial arrangement of collagen fibrils is the most fundamental ultrastructural characteristic of the normal general matrix. This, in turn, provides an efficient entrapment system for the swelling proteoglycans. Conversely, the most distinctive feature of the softened matrix is the presence of parallel and relatively unentwined fibrils, strongly aligned in the radial direction. The presence of an optically resolvable fibrous texture in the softened cartilage matrix indicates the presence of discrete bundles of closely packed and aligned fibrils at the ultrastructural level of organisation. The general absence of such texture in the normal cartilage general matrix is consistent with the much greater degree of interconnectedness and related short-range obliquity in the fibrillar architecture, hence the importance of the term pseudorandom network. A mechanism of structural transformation is proposed based on the important property of lateral interconnectivity in the fibrils which involves both entwinement and nonentwinement based interactions. The previously reported difference in intrinsic mechanical strength between the normal and softened matrices is consistent with the transformation model proposed in this study.

The chondrocyte and its pericellular microenvironment together represent the chondron, historically considered the primary structural, functional and metabolic unit of articular and other hyaline cartilages. This review summarises research over the last 10 years to establish the molecular anatomy, functional properties and metabolic contribution of the chondron in articular cartilage homeostasis, and its failure during the initiation and progression of degenerative osteoarthritis.

Cell–matrix and matrix–matrix interactions are of critical importance in regulating the development, maintenance and repair of articular cartilage. In this study, we examined the structural colocalisation of type VI collagen and fibronectin in isolated chondrons and long-term agarose cultured chondrocytes extracted from normal adult canine articular cartilage. Using double labelling immunohistochemistry in conjunction with dual channel confocal microscopy and digital image processing we demonstrate that type VI collagen and fibronectin are distributed in a similar staining pattern and are colocalised at the surface of cultured chondrocytes and isolated chondrons. The results suggest that type VI collagen and fibronectin may play a role in both cell–matrix adhesion and matrix–matrix cohesion in the pericellular microenvironment surrounding articular cartilage chondrocytes.