Assessment of a mechano-regulation theory of skeletal tissue differentiation in an in vivo model of mechanically induced cartilage formation

Mechanical cues are known to regulate tissue differentiation during skeletal healing. Quantitative characterization of this mechano-regulatory effect has great therapeutic potential. This study tested an existing theory that shear strain and interstitial fluid flow govern skeletal tissue differentiation by applying this theory to a scenario in which a bending motion applied to a healing transverse osteotomy results in cartilage, rather than bone, formation. A 3-D finite element mesh was created from micro-computed tomography images of a bending-stimulated callus and was used to estimate the mechanical conditions present in the callus during the mechanical stimulation. Predictions regarding the patterns of tissues—cartilage, fibrous tissue, and bone—that formed were made based on the distributions of fluid velocity and octahedral shear strain. These predictions were compared to histological sections obtained from a previous study. The mechano-regulation theory correctly predicted formation of large volumes of cartilage within the osteotomy gap and many, though not all patterns of tissue formation observed throughout the callus. The results support the concept that interstitial fluid velocity and tissue shear strain are key mec- hanical stimuli for the differentiation of skeletal tissues.     

Mechano-regulation of stem cell differentiation and tissue regeneration in osteochondral defects

Cartilage defects that penetrate the subchondral bone can undergo spontaneous repair through the formation of a fibrous or cartilaginous tissue mediated primarily by mesenchymal stem cells from the bone marrow. This tissue is biomechanically inferior to normal articular cartilage, and is often observed to degrade over time. Whether or not biomechanical factors control the type and quality of the repair tissue, and its subsequent degradation, have yet to be elucidated. In this paper, we hypothesise a relationship between the mechanical environment of mesenchymal stem cells and their subsequent dispersal, proliferation, differentiation and death. The mechano-regulation stimulus is hypothesised to be a function of strain and fluid flow; these quantities are calculated using biphasic poroelastic finite element analysis. A finite element model of an osteochondral defect in the knee was created, and used to simulate the spontaneous repair process. The model predicts bone formation through both endochondral and direct intramembranous ossification in the base of the defect, cartilage formation in the centre of the defect and fibrous tissue formation superficially. Greater amounts of fibrous tissue formation are predicted as the size of the defect is increased. Large strains are predicted within the fibrous tissue at the articular surface, resulting in significant cell apoptosis. This result leads to the conclusion that repair tissue degradation is initiated in the fibrous tissue that forms at the articular surface. The success of the mechano-regulation model in predicting many of the cellular events that occur during osteochondral defect healing suggest that in the future it could be used as a tool for optimising scaffolds for tissue engineering.     

A mechano-regulation model for tissue differentiation during fracture healing: analysis of gap size and loading

Bone has a capability to repair itself when it is fractured. Repair involves the generation of intermediate tissues, such as fibrous connective tissue, cartilage and woven bone, before final bone healing can occur. The intermediate tissues serve to stabilise the mechanical environment and provide a scaffold for differentiation of new tissues. The repair process is fundamentally affected by mechanical loading and by the geometric configuration of the fracture fragments. Biomechanical analyses of fracture healing have previously computed the stress distribution within the callus and identified the components of the stress tensor favouring or inhibiting differentiation of particular tissue phenotypes. In this paper, a biphasic poroelastic finite element model of a fracture callus is used to simulate the time-course of tissue differentiation during fracture healing. The simulation begins with granulation tissue (post-inflammation phase) and finishes with bone resorption. The biomechanical regulatory model assumes that tissue differentiation is controlled by a combination of shear strain and fluid flow acting within the tissue. High shear strain and fluid flows are assumed to deform the precursor cells stimulating formation of fibrous connective tissue, lower levels stimulate formation of cartilage, and lower again allows ossification. This mechano-regulatory scheme was tested by simulating healing in fractures with different gap sizes and loading magnitudes. The appearance and disappearance of the various tissues found in a callus was similar to histological observation. The effect of gap size and loading magnitude on the rate of reduction of the interfragmentary strain was sufficiently close to confirm the hypothesis that tissue differentiation phenomena could be governed by the proposed mechano-regulation model.     

Tissue engineering and cartilage

Human articular cartilage is an avascular structure, which, when injured, poses significant hurdles to repair strategies. Not only does the defect need to be repopulated with cells, but preferentially with hyaline-like cartilage.Successful tissue engineering relies on four specific criteria: cells, growth factors, scaffolds, and the mechanical environment. The cell population utilized may originate from cartilage itself (chondrocytes) or from growth factors that direct the development of mesenchymal stem cells toward a chondrogenic phenotype. These stem cells may originate from various mesenchymal tissues including bone marrow, synovium, adipose tissue, skeletal muscle, and periosteum. Another unique population of multipotent cells arises from Wharton''s jelly in human umbilical cords. A number of growth factors have been associated with chondrogenic differentiation of stem cells and the maintenance of the chondrogenic phenotype by chondrocytes in vitro, including TGFβ; BMP-2, 4 and 7; IGF-1; and GDF-5.Scaffolds chosen for effective tissue engineering with respect to cartilage repair can be protein based (collagen, fibrin, and gelatin), carbohydrate based (hyaluronan, agarose, alginate, PLLA/PGA, and chitosan), or formed by hydrogels. Mechanical compression, fluid-induced shear stress, and hydrostatic pressure are aspects of mechanical loading found in within the human knee joint, both during gait and at rest. Utilizing these factors may assist in stimulating the development of more robust cells for implantation.Effective tissue engineering has the potential to improve the quality of life of millions of patients and delay future medical costs related to joint arthroplasty and associated procedures.Key words: cartilage repair, gene therapy, growth factors, biomaterials, tissue engineering, stem cells, chondrocyte     

Tissue engineering and cartilage

Human articular cartilage is an avascular structure, which, when injured, poses significant hurdles to repair strategies. Not only does the defect need to be repopulated with cells, but preferentially with hyaline-like cartilage.

Successful tissue engineering relies on four specific criteria: cells, growth factors, scaffolds, and the mechanical environment. The cell population utilized may originate from cartilage itself (chondrocytes) or growth factors may direct the development of mesenchymal stem cells toward a chondrogenic phenotype. These stem cells may originate from various mesenchymal tissues including bone marrow, synovium, adipose tissue, skeletal muscle, and periosteum. Another unique population of multipotent cells arises from Wharton’s jelly in human umbilical cords. A number of growth factors have been associated with chondrogenic differentiation of stem cells and maintenance of the chondrogenic phenotype by chondrocytes in vitro, including TGF-β; BMP-2, 4, and 7; IGF-1; and GDF-5.

The scaffolds chosen for effective tissue engineering with respect to cartilage repair can be protein based (collagen, fibrin, and gelatin), carbohydrate based (hyaluronan, agarose, alginate, PLLA/PGA, and chitosan), or formed by hydrogels. Mechanical compression, fluid-induced shear stress, and hydrostatic pressure are all aspects of mechanical loading found in the human knee joint, both during gait and at rest. Utilizing these factors may assist in stimulating the development of more robust cells for implantation.

Effective tissue engineering has the ability to improve the quality of life of millions of patients and delay future medical costs related to joint arthroplasty and associated procedures.     

In silico study of bone tissue regeneration in an idealised porous hydrogel scaffold using a mechano-regulation algorithm

Mechanical stimulation, in the form of fluid perfusion or mechanical strain, enhances osteogenic differentiation and overall bone tissue formation by mesenchymal stems cells cultured in biomaterial scaffolds for tissue engineering applications. In silico techniques can be used to predict the mechanical environment within biomaterial scaffolds, and also the relationship between bone tissue regeneration and mechanical stimulation, and thereby inform conditions for bone tissue engineering experiments. In this study, we investigated bone tissue regeneration in an idealised hydrogel scaffold using a mechano-regulation model capable of predicting tissue differentiation, and specifically compared five loading cases, based on known experimental bioreactor regimes. These models predicted that low levels of mechanical loading, i.e. compression (0.5% strain), pore pressure of 10 kPa and a combination of compression (0.5%) and pore pressure (10 kPa), could induce more osteogenic differentiation and lead to the formation of a higher bone tissue fraction. In contrast greater volumes of cartilage and fibrous tissue fractions were predicted under higher levels of mechanical loading (i.e. compression strain of 5.0% and pore pressure of 100 kPa). The findings in this study may provide important information regarding the appropriate mechanical stimulation for in vitro bone tissue engineering experiments.     

A comparative microscopic and ultrastructural study of perichondrial tissue in cartilage of Octopus vulgaris (Cephalopoda, Mollusca)

The microscopic and submicroscopic structures of perichondrial tissues in the head cartilages of Octopus vulgaris were studied by polarized light and transmission electron microscopy. The orbital cartilages possess a birefringent layer parallel to the surface of the cartilage; ultrastructurally, this layer, which may be considered perichondrial tissue, has the typical organisation of connective tissue but does not possess the stratification of collagen laminae found in vertebrate perichondria. Perichondrial extracellular matrix is clearly distinct from that of cartilage because its collagen fibrils are of a larger diameter than collagen fibrils from cartilage. In addition, perichondrial fibroblasts are characteristically located at the center of collagen fibers. In the cerebral cartilage, the perichondrium is absent or discontinuous in relation to complex interconnections between cartilage and connective fibres, muscle fibres, blood vessels and nerve. Distinctive cartilage-lining cells, rich in electron dense cytoplasmatic granules, are stratified either along the cartilage surface or along vessels and muscle fibres that penetrate within the cartilage. The perichondrium of cephalopod cartilage, whose structure varies according to the location and function of its skeletal segments, mimics that of vertebrate perichondrium, exemplifying the high level of tissue differentiation attained by cephalopods.     

Rapid prototyping of anatomically shaped, tissue-engineered implants for restoring congruent articulating surfaces in small joints

Background:  Preliminary studies investigated advanced scaffold design and tissue engineering approaches towards restoring congruent articulating surfaces in small joints.
Materials and methods:  Anatomical femoral and tibial cartilage constructs, fabricated by three-dimensional fibre deposition (3DF) or compression moulding/particulate leaching (CM), were evaluated in vitro and in vivo in an autologous rabbit model. Effects of scaffold pore architecture on rabbit chondrocyte differentiation and mechanical properties were evaluated following in vitro culture and subcutaneous implantation in nude mice. After femoral and tibial osteotomy and autologous implantation of tissue-engineered constructs in rabbit knee joints, implant fixation and joint articulation were evaluated.
Results:  Rapid prototyping of 3DF architectures with 100% interconnecting pores promoted homogeneous distribution of viable cells, glycosaminoglycan (GAG) and collagen type II; significantly greater GAG content and differentiation capacity (GAG/DNA) in vitro compared to CM architectures; and higher mechanical equilibrium modulus and dynamic stiffness (at 0.1 Hz). Six weeks after implantation, femoral and tibial constructs had integrated with rabbit bone and knee flexion/extension and partial load bearing were regained. Histology demonstrated articulating surfaces between femoral and tibial constructs for CM and 3DF architectures; however, repair tissue appeared fibrocartilage-like and did not resemble implanted cartilage.
Conclusions:  Anatomically shaped, tissue-engineered constructs with designed mechanical properties and internal pore architectures may offer alternatives for reconstruction or restoration of congruent articulating surfaces in small joints.     

The restoration of full-thickness cartilage defects with mesenchymal stem cells (MSCs) loaded and cross-linked bilayer collagen scaffolds on rabbit model

Cartilage has a limited self-repair capability and the repair of large cartilage defects remains a challenge in clinic. This study aimed to investigate the effect of mesenchymal stem cells (MSCs) loaded three-dimensional bilayer collagen scaffold for cartilage repair. Cross-linked three-dimensional bilayer collagen scaffolds seeded with or without MSCs were implanted into large cartilage defects (4 mm in diameter; 3 mm in depth) in rabbit knees. The untreated cartilage defects served as control. The tissue response was evaluated at 6 and 12 weeks after implantation by general histology and semi-quantitative histological grading systems. In addition, the repaired tissues were evaluated by mechanical test at 12 weeks after implantation. The MSCs-loaded collagen scaffold group showed the most hyaline cartilage, highest histological scores and compressive modulus. Moreover, it showed a good integration with the subchondral bone and adjacent cartilage. The structure of the novel bilayer collagen scaffolds provided architectural support for the differentiation of MSCs and demonstrated successful induction of in vivo chondrogenesis. These findings suggested that MSCs-loaded bilayer collagen scaffold could be an appealing candidate to be used for cartilage regeneration.     

Collagen IX is indispensable for timely maturation of cartilage during fracture repair in mice.

Fracture repair recapitulates in adult organisms the sequence of cell biological events of endochondral ossification during skeletal development and growth. After initial inflammation and deposition of granulation tissue, a cartilaginous callus is formed which, subsequently, is remodeled into bone. In part, bone formation is influenced also by the properties of the extracellular matrix of the cartilaginous callus. Deletion of individual macromolecular components can alter extracellular matrix suprastructures, and hence stability and organization of mesenchymal tissues. Here, we took advantage of the collagen IX knockout mouse model to better understand the role of this collagen for organization, differentiation and maturation of a cartilaginous template during formation of new bone. Although a seemingly crucial component of cartilage fibrils is missing, collagen IX-deficient mice develop normally, but are predisposed to premature joint cartilage degeneration. However, we show here that lack of collagen IX alters the time course of callus differentiation during bone fracture healing. The maturation of cartilage matrix was delayed in collagen IX-deficient mice calli as judged by collagen X expression during the repair phase and the total amount of cartilage matrix was reduced. Entering the remodeling phase of fracture healing, Col9a1(-/-) calli retained a larger percentage of cartilage matrix than in wild type indicating also a delayed formation of new bone. We concluded that endochondral bone formation can occur in collagen IX knockout mice but is impaired under conditions of stress, such as the repair of an unfixed fractured long bone.     

Remodelling of collagen fibre transition stretch and angular distribution in soft biological tissues and cell-seeded hydrogels

The extracellular matrix in many biological tissues is adapted to its mechanical environment. In this study, a phenomenological model for collagen remodelling is introduced that incorporates angular remodelling (fibre reorientation) and the adaptation of the so-called transition stretch. This is achieved by introducing a local stress-free configuration for the collagen network by a multiplicative decomposition of the deformation gradient and the appropriate definition of the anisotropic free Helmholtz energy potentials and structure tensors. The collagen network is either treated using discrete fibre directions or a continuous angular distribution. The first part of the study illustrates the influence of force- and displacement-controlled loading on either stress- or deformation-driven remodelling processes in tissues with various degrees of fibre reinforcement. The model is then applied to recent experimental studies of collagen remodelling, specifically periosteum adaptation (Foolen et?al. in J Biomech 43(16):3168–3176, 2010), collagen gel (Thomopoulos et?al. in J Biomech Eng 127(5):742–750, 2005) and fibrin cruciform (Sander et?al. in Ann Biomed Eng 1–16, 2010) compaction. The model is able to capture the basic effects of an adapting transition stretch over time in the periosteal simulations, as well as the compaction and the development of structural anisotropy in the collagen and fibrin gels. The model can potentially be applied to elucidate structure–function relationships, better interpret in vitro experiments involving collagen remodelling, and help investigate aspects of certain pathologies, such as connective tissue contracture.     

Comparison of biophysical stimuli for mechano-regulation of tissue differentiation during fracture healing

Most long-bone fractures heal through indirect or secondary fracture healing, a complex process in which endochondral ossification is an essential part and bone is regenerated by tissue differentiation. This process is sensitive to the mechanical environment, and several authors have proposed mechano-regulation algorithms to describe it using strain, pore pressure and/or interstitial fluid velocity as biofeedback variables. The aim of this study was to compare various mechano-regulation algorithms' abilities to describe normal fracture healing in one computational model. Additionally, we hypothesized that tissue differentiation during normal fracture healing could be equally well regulated by the individual mechanical stimuli, e.g. deviatoric strain, pore pressure or fluid velocity. A biphasic finite element model of an ovine tibia with a 3mm fracture gap and callus was used to simulate the course of tissue differentiation during normal fracture healing. The load applied was regulated in a biofeedback loop, where the load magnitude was determined by the interfragmentary movement in the fracture gap. All the previously published mechano-regulation algorithms studied, simulated the course of normal fracture healing correctly. They predicted (1) intramembranous bone formation along the periosteum and callus tip, (2) endochondral ossification within the external callus and cortical gap, and (3) creeping substitution of bone towards the gap from the initial lateral osseous bridge. Some differences between the effects of the algorithms were seen, but they were not significant. None of the volumetric components, i.e. pore pressure or fluid velocity, alone were able to correctly predict spatial or temporal tissue distribution during fracture healing. However, simulation as a function of only deviatoric strain accurately predicted the course of normal fracture healing. This suggests that the deviatoric component may be the most significant mechanical parameter to guide tissue differentiation during indirect fracture healing.     

Time-dependent functional maturation of scaffold-free cartilage tissue analogs

One of the most critical parameters in cartilage tissue engineering which influences the clinical success of a repair therapy is the ability to match the load-bearing capacity of the tissue as it functions in vivo. While mechanical forces are known to positively influence the development of cartilage matrix architecture, these same forces can induce long-term implant failure due to poor integration or structural deficiencies. As such, in the design of optimal repair strategies, it is critical to understand the timeline of construct maturation and how the elaboration of matrix correlates with the development of mechanical properties. We have previously characterized a scaffold-free method to engineer cartilage utilizing primary chondrocytes cultured at high density in hydrogel-coated culture vessels to promote the formation of a self-aggregating cell suspension that condenses to form a cartilage-like biomass, or cartilage tissue analog (CTA). Chondrocytes in these CTAs maintain their cellular phenotype and deposit extracellular matrix to form a construct that has characteristics similar to native cartilage; however, the mechanical integrity of CTAs had not yet been evaluated. In this study, we found that chondrocytes within CTAs produced a robust matrix of proteoglycans and collagen that correlated with increasing mechanical properties and decreasing cell-matrix ratios, leading to properties that approached that of native cartilage. These results demonstrate a unique approach to generating a cartilage-like tissue without the complicating factor of scaffold, while showing increased compressive properties and matrix characteristics consistent with other approaches, including scaffold-based constructs. To further improve the mechanics of CTAs, studies are currently underway to explore the effect of hydrodynamic loading and whether these changes would be reflective of in vivo maturation in animal models. The functional maturation of cartilage tissue analogs as described here support this engineered cartilage model for use in clinical and experimental applications for repair and regeneration in joint-related pathologies.     

The use of small angle light scattering in assessing strain induced collagen degradation in arterial tissue ex vivo

Collagen is the predominant load bearing component in many soft tissues including arterial tissue and is therefore critical in determining the mechanical integrity of such tissues. Degradation of collagen fibres is hypothesized to be a strain dependent process whereby the rate of degradation is affected by the magnitude of strain applied to the collagen fibres. The aim of this study is to investigate the ability of small angle light scattering (SALS) imaging to identify strain dependent degradation of collagen fibres in arterial tissue ex vivo, and determine whether a strain induced protection mechanism exists in arterial tissue as observed in pure collagen and other collagenous tissues. SALS was used in combination with histological and second harmonic generation (SHG) analysis to determine the collagen fibre architecture in arterial tissue subjected to strain directed degradation. SALS alignment analysis identified statistically significant differences in fibre alignment depending on the strain magnitude applied to the tissue. These results were also observed using histology and SHG. Our findings suggest a strain protection mechanism may exist for arterial collagen at intermediate strain magnitudes between 0% and 25%. These findings may have implications for the onset and progression of arterial disease where changes in the mechanical environment of arterial tissue may lead to changes in the collagen degradation rate.     

Type III collagen is a key regulator of the collagen fibrillar structure and biomechanics of articular cartilage and meniscus

Despite the fact that type III collagen is the second most abundant collagen type in the body, its contribution to the physiologic maintenance and repair of skeletal tissues remains poorly understood. This study queried the role of type III collagen in the structure and biomechanical functions of two structurally distinctive tissues in the knee joint, type II collagen-rich articular cartilage and type I collagen-dominated meniscus. Integrating outcomes from atomic force microscopy-based nanomechanical tests, collagen fibril nanostructural analysis, collagen cross-link analysis and histology, we elucidated the impact of type III collagen haplodeficiency on the morphology, nanostructure and biomechanical properties of articular cartilage and meniscus in Col3a1^+/− mice. Reduction of type III collagen leads to increased heterogeneity and mean thickness of collagen fibril diameter, as well as reduced modulus in both tissues, and these effects became more pronounced with skeletal maturation. These data suggest a crucial role of type III collagen in mediating fibril assembly and biomechanical functions of both articular cartilage and meniscus during post-natal growth. In articular cartilage, type III collagen has a marked contribution to the micromechanics of the pericellular matrix, indicating a potential role in mediating the early stage of type II collagen fibrillogenesis and chondrocyte mechanotransduction. In both tissues, reduction of type III collagen leads to decrease in tissue modulus despite the increase in collagen cross-linking. This suggests that the disruption of matrix structure due to type III collagen deficiency outweighs the stiffening of collagen fibrils by increased cross-linking, leading to a net negative impact on tissue modulus. Collectively, this study is the first to highlight the crucial structural role of type III collagen in both articular cartilage and meniscus extracellular matrices. We expect these results to expand our understanding of type III collagen across various tissue types, and to uncover critical molecular components of the microniche for regenerative strategies targeting articular cartilage and meniscus repair.     

Calcification or dedifferentiation: Requirement to lock mesenchymal stem cells in a desired differentiation stage

A current challenge in mesenchymal stem cell (MSC)‐based cartilage repair is to solve donor and tissue‐dependent variability of MSC cultures and to prevent chondrogenic cells from terminal differentiation like in the growth plate. The aim of this study was to select the best source for MSC which could promise stable cartilage formation in the absence of hypertrophy and ectopic in vivo mineralization. We hypothesized that MSC from synovium are superior to bone marrow‐ and adipose tissue‐derived MSC since they are derived from a joint tissue. MSC were characterized by flow cytometry. MSC pellets were cultured under chondrogenic conditions and differentiation was evaluated by histology, gene expression analysis, and determination of alkaline phosphatase activity (ALP). After chondrogenic induction, pellets were transplanted subcutaneously into SCID mice. MSC from bone marrow, adipose tissue, and synovium revealed similar COL2A1/COL10A1 mRNA levels after chondrogenic induction and were positive for collagen‐type‐X. Bone marrow‐derived and adipose tissue‐derived MSC showed significantly higher ALP activity than MSC from synovium. Low ALP‐activity before transplantation of pellets correlated with marginal calcification of explants. Surprisingly, non‐mineralizing transplants specifically lost their collagen‐type II, but not collagen‐type I deposition in vivo, or were fully degraded. In conclusion, the lower donor‐dependent ALP activation and reduced mineralization of synovium‐derived heterotopic transplants did not lead to stable ectopic cartilage as known from articular chondrocytes, but correlated with fibrous dedifferentation or complete degeneration of MSC pellets. This emphasizes that beside appropriate induction of differentiation, locking of MSC in the desired differentiation state is a major challenge for MSC‐based repair strategies. J. Cell. Physiol. 219: 219–226, 2009. © 2008 Wiley‐Liss, Inc.     

Potential use of the human amniotic membrane as a scaffold in human articular cartilage repair

The human amniotic membrane (HAM) is an abundant and readily obtained tissue that may be an important source of scaffold for transplanted chondrocytes in cartilage regeneration in vivo. To evaluate the potential use of cryopreserved HAMs as a support system for human chondrocytes in human articular cartilage repair. Chondrocytes were isolated from human articular cartilage, cultured and grown on the chorionic basement membrane side of HAMs. HAMs with chondrocytes were then used in 44 in vitro human osteoarthritis cartilage repair trials. Repair was evaluated at 4, 8 and 16 weeks by histological analysis. Chondrocytes cultured on the HAM revealed that cells grew on the chorionic basement membrane layer, but not on the epithelial side. Chondrocytes grown on the chorionic side of the HAM express type II collagen but not type I, indicating that after being in culture for 3–4 weeks they had not de-differentiated into fibroblasts. In vitro repair experiments showed formation on OA cartilage of new tissue expressing type II collagen. Integration of the new tissue with OA cartilage was excellent. The results indicate that cryopreserved HAMs can be used to support chondrocyte proliferation for transplantation therapy to repair OA cartilage.     

Alterations in structure and properties of collagen network of osteoarthritic and repaired cartilage modify knee joint stresses

Organization of the collagen network is known to be different in healthy, osteoarthritic and repaired cartilage. The aim of the study was to investigate how the structure and properties of collagen network of cartilage modulate stresses in a knee joint with osteoarthritis or cartilage repair. Magnetic resonance imaging (MRI) at 1.5 T was conducted for a knee joint of a male subject. Articular cartilage and menisci in the knee joint were segmented, and a finite element mesh was constructed based on the two-dimensional section in sagittal projection. Then, the knee joint stresses were simulated under impact loads by implementing the structure and properties of healthy, osteoarthritic and repaired cartilage in the models. During the progression of osteoarthritis, characterized especially by the progressive increase in the collagen fibrillation from the superficial to the deeper layers, the stresses were reduced in the superficial zone of cartilage, while they were increased in and under menisci. Increased fibril network stiffness of repair tissue with randomly organized collagen fibril network reduced the peak stresses in the adjacent tissue and strains at the repair–adjacent cartilage interface. High collagen fibril strains were indicative of stress concentration areas in osteoarthritic and repaired cartilage. The collagen network orientation and stiffness controlled the stress distributions in healthy, osteoarthritic and repaired cartilage. The evaluation of articular cartilage function using clinical MRI and biomechanical modeling could enable noninvasive estimation of osteoarthritis progression and monitoring of cartilage repair. This study presents a step toward those goals.     

Three dimensional patient-specific collagen architecture modulates cartilage responses in the knee joint during gait

Site-specific variation of collagen fibril orientations can affect cartilage stresses in knee joints. However, this has not been confirmed by 3-D analyses. Therefore, we present a novel method for evaluation of the effect of patient-specific collagen architecture on time-dependent mechanical responses of knee joint cartilage during gait. 3-D finite element (FE) models of a human knee joint were created with the collagen architectures obtained from T₂ mapped MRI (patient-specific model) and from literature (literature model). The effect of accuracy of the implementation of collagen fibril architecture into the model was examined by using a submodel with denser FE mesh. Compared to the literature model, fibril strains and maximum principal stresses were reduced especially in the superficial/middle regions of medial tibial cartilage in the patient-specific model after the loading response of gait (up to ?413 and ?26%, respectively). Compared to the more coarsely meshed joint model, the patient-specific submodel demonstrated similar strain and stress distributions but increased values particularly in the superficial cartilage regions (especially stresses increased >60%). The results demonstrate that implementation of subject-specific collagen architecture of cartilage in 3-D modulates location- and time-dependent mechanical responses of human knee joint cartilage. Submodeling with more accurate implementation of collagen fibril architecture alters cartilage stresses particularly in the superficial/middle tissue.     

The role of tissue interactions in the skeletogenic differentiation of avian neural crest cells

In the avian embryo, cranial neural crest (NC) cells migrate extensively throughout the head region and give rise to most of the cranial skeleton (Le Lievre, C. S. (1978). J. Embryol. Exp. Morphol.47, 17–37). To investigate the skeletogenic differentiation of these cells, NC explants from the mesencephalic level of st. 9+ embryos were grown in standard organ culture on Millipore filter substrates either in isolation or in combination with those tissues with which the cells normally associate during their in vivo migration and at their final tissue sites. The results demonstrate that interaction between premigratory NC and cranial ectoderm leads to chondrogenic differentiation of NC cells. Combination of premigratory NC with presumptive site tissues led to a pattern of NC cell differentiation normally expressed after in vivo migration: Combinations of NC with retinal pigmented epithelia gave cartilage, whereas NC with maxillary ectoderm formed cartilage and membrane bone. Both resulting skeletal tissues possessed their characteristic collagen types (II in cartilage and I in bone) as shown by indirect immunofluorescence using antibodies raised against specific types of collagen. It is concluded that avian cephalic NC cells require tissue interactions if chondrogenic and osteogenic differentiation is to ensue, but that migration per se is not an absolute prerequisite for these types of differentiation. The degree of specificity underlying such interactions is discussed.