Marcy Wong

Articular cartilage functional histomorphology and mechanobiology: a research perspective

The histomorphogenesis of articular cartilage is regulated during skeletal development by the intermittent forces and motions imposed at diarthrodial joints. A key feature in this development is the formation of the superficial, transitional, radial, and calcified cartilage zones through the cartilage thickness. The histomorphological, biological, and mechanical characteristics of these zones can be correlated with the distributions of pressures, deformations, and pressure-induced fluid flow that are created in vivo. In a mature joint, cyclic loads produce cyclic hydrostatic fluid pressure through the entire cartilage thickness that is comparable in magnitude to the applied joint pressure. Prolonged physical activity can cause the total cartilage thickness to decrease about 5%, although the consolidation strains vary tremendously in the different zones. The superficial zone can experience significant fluid exudation and consolidation (compressive strains) in the range of 60% while the radial zone experiences relatively little fluid flow and consolidation. The topological variation in the histomorphologic appearance of articular cartilage is influenced by the local mechanical loading of chondrocytes in the different zones. Patterns of stress, strain, and fluid flow created in the joint result in spatial and temporal changes in the rates of synthesis and degradation of matrix proteins. When viewed over the course of a lifetime, even subtle difference in these cellular processes can affect the micro- and macro-morphology of articular cartilage. This hypothesis is supported by in vivo and ex vivo experiments where load-induced changes in matrix synthesis and catabolism, gene expression, and signal transduction pathways have been observed.

Cyclic compression of articular cartilage explants is associated with progressive consolidation and altered expression pattern of extracellular matrix proteins

In this study, we investigated the biosynthetic response of full thickness, adult bovine articular cartilage explants to 45 h of static and cyclic unconfined compression. The cyclic compression of articular cartilage resulted in a progressive consolidation of the cartilage matrix. The oscillatory loading increased protein synthesis ([35S]methionine incorporation) by as much as 50% above free swelling control values, but had an inhibitory influence on proteoglycan synthesis ([35SO4] incorporation). As expected, static compression was associated with a dose-dependent decrease in biosynthetic activity. ECM oligomeric proteins which were most affected by mechanical loading were fibronectin and cartilage oligomeric matrix protein (COMP). Static compression at all amplitudes caused a significant increase in fibronectin synthesis over free swelling control levels. Cyclic compression of articular cartilage at 0.1 Hz and higher was consistently associated with a dramatic increase in the synthesis of COMP as well as fibronectin. The biosynthetic activity of chondrocytes appears to be sensitive to both the frequency and amplitude of the applied load. The results of this study support the hypothesis that cartilage tissue can remodel its extracellular matrix in response to alterations in functional demand.

Volumetric changes of articular cartilage during stress relaxation in unconfined compression.

The time-dependent lateral expansion and load relaxation of cartilage cylinders subjected to unconfined compression were simultaneously recorded. These measurements were used to (1) test the assumption of incompressibility for articular cartilage, (2) measure the Poissons ratio of articular cartilage in compression and (3) investigate the relationship between stress relaxation and volumetric change. Mechanical tests were performed on fetal, calf, and adult humeral head articular cartilage. The instantaneous Poissons ratio of adult cartilage was 0.49+/-0.08 (mean+S.D.), thus confirming the assumption of incompressibility for this tissue. The instantaneous Poissons ratio was significantly lower for calf (0. 38+/-0.04) and fetal cartilage (0.36+/-0.04). The equilibrium Poissons ratio, i.e. true Poissons ratio of the solid matrix, was significantly higher for the adult tissue (0.26+/-0.11) compared to both the fetal (0.09+/-0.02) and calf (0.11+/-0.03) cartilage. A linear relationship between time-matched load and lateral expansion after the first minute of stress relaxation was observed.

Development of mechanically stable alginate/chondrocyte constructs: effects of guluronic acid content and matrix synthesis

The purpose of this study was to investigate factors which enhanced the compressive properties of alginate/chondrocyte constructs. Firstly, we studied the effect of biochemical composition (high, mid and low guluronic acid content) and sterilization method on alginate properties. Secondly, we studied the biosynthetic characteristics of chondrocytes in three different alginate compositions and performed mechanical tests to determine whether the synthesis of cartilage matrix components could significantly enhance the compressive properties. 2% alginate solutions containing an initial cell density of 4 × 106 cells/ml were cast into cylinders and cultured for seven weeks. Compression tests, biochemistry, immunohistochemistry and electron microscopy were performed at fixed intervals during the seven‐week culture period. The dynamic modulus, peak strain, and peak stress were maximum for alginate with the highest guluronic acid content. The presence of cells and their respective matrix components enhanced the equilibrium modulus of the constructs for all types of alginate, though this effect was small. Alginate containing the middle amount of guluronic acid resulted in constructs which were both mechanically stable and which promoted synthesis of cartilage matrix proteins. In experiments and applications in which the mechanical integrity of the alginate is important, the composition and purity of the alginate and its method of sterilization should be selected with care.

Collagen Fibrillogenesis by Chondrocytes in Alginate

Collagen is the primary structural component in connective tissue. The poor mechanical properties of most cell-seeded cartilage grafts used for cartilage repair can be attributed to the low level of collagen synthesized compared with native cartilage. In this study, the synthesis and assembly of collagen by chondrocytes in hydrogels were investigated, with particular attention paid to the role of cross-link formation in this process. Primary bovine chondrocytes were seeded in alginate and collagen synthesis was assessed in the presence and absence of beta-aminopropronitrile (BAPN), a potent inhibitor of the enzyme lysyl oxidase and collagen cross-link formation. Cultures on days 21, 35, and 49 were evaluated by stereology, biochemistry, and real-time reverse transcriptase-polymerase chain reaction. All measures of collagen synthesis (except hydroxyproline) significantly increased in the presence of 0.25 mM BAPN. By 35 days of culture, the average collagen fibril diameter was 62 +/- 10 nm in control cultures and 109 +/- 20 nm with BAPN supplementation. The collagen volume density increased from 5 +/- 3% in control cultures to 17 +/- 1% in the presence of BAPN. Likewise, the expression of cartilage-specific collagens (type II and XI) and aggrecan increased significantly as a result of BAPN culture. These findings demonstrate the prominent role of collagen cross-linking in collagen fibrillogenesis and suggest approaches by which collagen synthesis and assembly could be controlled in tissue-engineered constructs.

Articular Cartilage Biology and Biomechanics

The intricate patterns of movement seen in vertebrate animals are made possible through the elegant design of synovial joints. The synovial joint is an organ whose function depends on several connective tissues including bone, ligament, tendon, synovium and articular cartilage. Of these, articular cartilage has perhaps the most highly specialized biomechanical properties. It provides the lubricated bearing surface which permits skeletal elements to glide and rotate against each other in a friction-free manner. In addition, the superficial layers of articular cartilage act as a deformable cushion which distributes and attenuates the peak loads associated with physical activity. Articular cartilage is also an extremely resilient tissue which can withstand tens of millions of cycles of load over the course of its lifetime.

Influence of tissue shear deformation on chondrocyte biosynthesis and matrix nano-electromechanics

Articular cartilage provides lubrication and load bearing functions during the motion of synovial joints. Such a specialized biomechanical function is enabled by the mechanical and electromechanical properties of cartilage extracellular matrix (ECM) and the interaction between cartilage and synovial fluid. Within cartilage matrix, highly charged aggrecan molecules are embedded within a dense collagen fibrillar network. The proteoglycan-associated repulsive forces are restrained by tensile forces within the collagen network. At the molecular level, these repulsive or swelling stresses are mostly due to electrical double layer repulsion associated with the negative fixed charges on glycosaminoglycan (GAG) chains, in addition to the elastic and entropic interactions between GAG macromolecules.