The mechanical influence of bone spicules in the osteochondral junction: A finite element modelling study

Biomechanics and Modeling in Mechanobiology - While much has been done to study how cartilage responds to mechanical loading, as well as modelling such responses, arguably less has been...     

A new device to control mechanical environment in bone defect healing in rats

Mechanical conditions have a significant influence on the biological processes of bone healing. Small animal models that allow controlling the mechanical environment of fracture and bone defect healing are needed. The aim of this study was to develop a new animal model that allows to reliably control the mechanical environment in fracture and bone defect healing in rats using different implant materials. An external fixator was designed and mounted in vitro to rat femurs using four Kirschner-wires (titanium (T) or steel (S)) of 1.2mm diameter. The specimens were distracted to a gap of 1.5mm. Axial and torsional stiffness of the device was tested increasing the offset (distance between bone and fixator crossbar) from 5 to 15mm. In vivo performance (well-being, infection, breaking of wires and bone healing) was evaluated in four groups of 24 Sprague-Dawley rats varying in offset (7.5 and 15mm) and implant material (S/T) over 6 weeks. Torsional and axial stiffness were higher in steel compared to titanium setups. A decrease in all configurations was observed by increasing the offset. The offset 7.5mm showed a significantly higher torsional (S: p<0.01, T: p<0.001) and axial in vitro stiffness (S: p<0.001, T: p<0.001) compared to 15mm offset of the fixator. Although in vitro designed to be different in mechanical stiffness, no difference was found between the groups regarding complication rate. The overall-complication rate was 5.2%. In conclusion, we were able to establish a small animal model for bone defect healing which allows modeling the mechanical conditions at the defect site in a defined manner.     

Differentiation of transplanted mesenchymal stem cells in a large osteochondral defect in rabbit

BACKGROUND: Although accumulating evidence shows that mesenchymal stem cells (MSC) are a promising cell source for articular cartilage repair, the fate of transplanted MSC has not been extensively studied. METHODS: To monitor their persistence and differentiation, we labeled uninduced MSC with a fluorescent dye, PKH26, and transplanted them, in a poly-glycolic-acid scaffold, to full-thickness defects made in the weight-bearing area of rabbit femoral trochleae with hyaluronate sheets. The fate of the labeled cells was monitored for up to 8 weeks. RESULTS: Two weeks after transplantation, immature cartilage containing collagen type II had formed. By 8 weeks, this cartilage had thinned and immunolabeling for collagen type II gradually disappeared from the basal region, which became positive for collagen type I. Most chondrocytes within the regenerated cartilage were PKH26-positive and, therefore, derived from transplanted MSC, whereas osteoblasts within the regenerated bone were a mixture of donor- and host-derived cells. The thickness of the cartilage became thinner up to 8 weeks and then remained stable up to 42 weeks after surgery. DISCUSSION: These results showed that uninduced MSC were able to survive osteochondral defects and differentiated according to the environment, making a major contribution to initial cartilage formation and a partial contribution to bone regeneration.     

The influence of test conditions on characterization of the mechanical properties of brain tissue

To understand brain injuries better, the mechanical properties of brain tissue have been studied for 50 years; however, no universally accepted data set exists. The variation in material properties reported may be caused by differences in testing methods and protocols used. An overview of studies on the mechanical properties of brain tissue is given, focusing on testing methods. Moreover, the influence of important test conditions, such as temperature, anisotropy, and precompression was experimentally determined for shear deformation. The results measured at room temperature show a stiffer response than those measured at body temperature. By applying the time-temperature superposition, a horizontal shift factor a(T)=8.5-11 was found, which is in agreement with the values found in literature. Anisotropy of samples from the corona radiata was investigated by measuring the shear resistance for different directions in the sagittal, the coronal, and the transverse plane. The results measured in the coronal and the transverse plane were 1.3 and 1.25 times stiffer than the results obtained from the sagittal plane. The variation caused by anisotropy within the same plane of individual samples was found to range from 25% to 54%. The effect of precompression on shear results was investigated and was found to stiffen the sample response. Combinations of these and other factors (postmortem time, donor age, donor type, etc.) lead to large differences among different studies, depending on the different test conditions.     

Effects of mechanical loading patterns,bone graft,and proximity to periosteum on bone defect healing

The goal of this study is to elucidate whether mechanobiological factors, including mechanical loading patterns, presence of bone graft, and proximity to the periosteum, correlate to de novo tissue generation and healing in critical sized long bone defects, which are enveloped by periosteum in situ and are bridged at 16 weeks, in sheep femora. Quantitative histomorphometric measures of defect cross sections show that, along the axis least able to resist bending loads (minor centroidal axis, CA), bone laid down in the first two weeks after surgery exhibits more mineralization albeit less total area compared to bone along the axis most able to resist bending loads (major CA). Similarly, areas of the cross section along the minor CA show a higher degree of perfusion albeit less total area of perfusion compared to bone along the major CA. Furthermore, proximity to the periosteal niche, in conjunction with the presence of bone graft and predominant loading patterns, relates significantly to the radial distribution of early bone apposition and perfusion of bone at 16 weeks after surgery (linear regression with R²>0.80). In the absence of graft, early bone density is relatively evenly distributed in the defect zone, as is the intensity of perfused tissue. As measured by a steeper average slope in intensity of fluorochrome (new bone) distribution between the periosteum and the IM nail, the presence of bone graft retards initial bone formation in the defect zone and is associated with less evenly distributed tissue perfusion (steeper slope) persisting 16 weeks after surgery. Finally, although the mean area of bone resorption is not significantly different within or between groups defined by the presence of graft and/or mechanical loading patterns in the defect zone, the mean area of infilling resorption spaces is significantly higher in areas of the defect zone least able to resist bending (minor CA) but is not significantly related to the presence of bone graft. To our knowledge, the use of the major and minor centroidal axes to relate prevailing mechanical loading patterns to area and density of early bone generation in bone defects has not been reported prior to this study and may provide a new means to assess structure–function relationships in de novo bone generation and healing of bone defects.     

Influence of the stiffness of bone defect implants on the mechanical conditions at the interface--a finite element analysis with contact

The study focused on the influence of the implant material stiffness on stress distribution and micromotion at the interface of bone defect implants. We hypothesized that a low-stiffness implant with a modulus closer to that of the surrounding trabecular bone would yield a more homogeneous stress distribution and less micromotion at the interface with the bony bed. To prove this hypothesis we generated a three-dimensional, non-linear, anisotropic finite element (FE) model. The FE model corresponded to a previously developed animal model in sheep. A prismatic implant filled a standardized defect in the load-bearing area of the trabecular bone beneath the tibial plateau. The interface was described by face-to-face contact elements, which allow press fits, friction, sliding, and gapping. We assumed a physiological load condition and calculated contact pressures, shear stresses, and shear movements at the interface for two implants of different stiffness (titanium: E=110GPa; composite: E=2.2GPa). The FE model showed that the stress distribution was more homogeneous for the low-stiffness implant. The maximum pressure for the composite implant (2.1 MPa) was lower than for the titanium implant (5.6 MPa). Contrary to our hypothesis, we found more micromotion for the composite (up to 6 microm) than for the titanium implant (up to 4.5 microm). However, for both implants peak stresses and micromotion were in a range that predicts adequate conditions for the osseointegration. This was confirmed by the histological results from the animal studies.     

Fibroblasts change spreading capability and mechanical properties in a direct interaction with keratinocytes in conditions mimicking wound healing

Keratinocytes are predominant in the uppermost layer of the skin, while fibroblasts dominate in the dermal layer. These cells interact with each other directly when fibroblasts migrate to a region of the wound where they induce keratinocytes proliferation through double paracrine signalling. Since a response from both keratinocytes and fibroblasts dominates during the inflammatory and proliferative phases, the exact knowledge how these two types of cells interact with each other is crucial for deeper understanding of mechanisms involved in the wound healing process. The aim of this study was to quantify alterations in mechanical properties of cells, i.e. fibroblasts and keratinocytes, in conditions mimicking direct cellular interactions observed in wound healing. Single cell elasticity was measured using atomic force microscope. To verify the influence of keratinocyte neighbors on fibroblasts elasticity (and vice versa), the effect of cellular confluency was studied in parallel. Our results enabled us to distinguish cellular density-related effects from intercellular interactions occurring between fibroblasts and keratinocytes. While the presence of keratinocytes affects fibroblasts spreading capability and mechanical properties, the keratinocytes remain unaffected by the fibroblasts. These results highlight the importance of the cellular deformability in understanding of the role of biomechanics in double paracrine signalling as fibroblast-keratinocyte interaction can change the potential of the wound healing.     

The influence of doubly attached crossbridges on the mechanical behavior of skeletal muscle fibers under equilibrium conditions.

A simple model of a double-headed crossbridge is introduced to explain the retardation of force decay after an imposed stretch in skeletal muscle fibers under equilibrium conditions. The critical assumption in the model is that once one of the heads of a crossbridge is attached to one of the available actin sites, the attachment of the second head will be restricted to a level of strain determined by the attachment of the first head. The crossbridge structure, namely the connection of both heads of a crossbridge to the same tail region, is assumed to impose this constraint on the spatial configurations of crossbridge heads. The unique feature of the model is the prediction that, in the presence of a ligand (PPi, ADP, AMP-PNP) and absence of Ca2+, the halftime of force decay is many times larger than the inverse rate of detachment of a crossbridge head measured in solution. This prediction is in agreement with measured values of half-times of force decay in fibers under similar conditions (Schoenberg, M., and E. Eisenberg. 1985. Biophys. J. 48:863-871f). It is predicted that a crossbridge head is more likely to re-attach to its previously strained position than remain unattached while the other head is attached, leading to the slow decay of force. Our computations also show that the apparent cooperativity in crossbridge binding observed in experiments (Brenner, B., L. C. Yu, L. E. Greene, E. Eisenberg, and M. Schoenberg. 1986. Biophys. J. 50:1101-1108) can be partially accounted by the double-headed crossbridge attachment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Indentation of an osteochondral repair: sensitivity to experimental variables and boundary conditions

The sensitivity of the affects of indenter radius, defect depth, cartilage permeability and flow boundary conditions, on the indentation testing of a repairing osteochondral defect was investigated. Since the boundary condition on the flow across the cartilage repair-subchondral bone interface is not known, the effects of two different conditions were investigated: free-flow and no-flow. A poroelastic finite element model of an osteochondral defect at different stages of the repair process was developed using dimensions typical of the rabbit knee. Results showed when the radius of the indenter was much less than the thickness of the cartilage the sensitivity of the indentation displacement to flow boundary conditions decreased. Simulated indentation displacement was insensitive to bone regeneration up to 50% of the initial defect depth, which suggests that only the properties of the material in the upper-half of the defect are being evaluated. Small variations in permeability had little affect on the simulated indentation. In a fully repaired defect, the simulated indentation is independent of the boundary condition. However, while the defect is in the process of repair and the regenerated cartilage is deeper than the host, indentation is sensitive to the flow boundary condition. Based on these results, and feasible experimental conditions, we conclude that the boundary condition on the repair-subchondral bone interface must be known in all cases except when the defect approaches full repair, if accurate estimates of material properties are to be obtained from indentation.     

Behavior of transplanted bone marrow-derived GFP mesenchymal cells in osteochondral defect as a simulation of autologous transplantation.

To elucidate the behavior of autologously transplanted mesenchymal cells in osteochondral defects, we followed transplanted cells using green fluorescent protein (GFP) transgenic rats, in which all cells express GFP signals in their cytoplasm and nuclei as transplantation donors. Bone marrow-derived mesenchymal cells, which contain mesenchymal stem cells (MSCs), were obtained from transgenic rats. Then, dense mesenchymal cell masses created by hanging-drop culture were transplanted and fixed with fibrin glue into osteochondral defects of wild-type rats. At 24 weeks after surgery, the defects were repaired with hyaline-like cartilage and subchondral bone. GFP positive cells, indicating transplanted mesenchymal-derived cells, were observed in the regenerated tissues for 24 weeks although GFP positive cells decreased in number with time. Because GFP causes no immunological rejection and requires no chemicals for visualization, transplantation between transgenic and wild-type rats can be regarded as a simulation of autologous transplantation, and the survivability of transplanted cells are able to be followed easily and reliably. Thus, the behavior of transplanted mesenchymal cells was able to be elucidated in vivo by this strategy, and the results could be essential in future tissue engineering for the regeneration of osteochondral defects with original hyaline cartilage and subchondral bone.     

Evaluating osteochondral defect repair potential of autologous rabbit bone marrow cells on type II collagen scaffold

The feasibility of using genipin cross-linked type II collagen scaffold with rabbit bone marrow mesenchymal stem cells (RBMSCs) to repair cartilage defect was herein studied. Induction of RBMSCs into chondrocytic phenotype on type II collagen scaffold in vitro was conducted using TGF-β 3 containing medium. After 3-weeks of induction, chondrocytic behavior, including marker genes expression and specific extracellular matrix (ECM) secretion, was observed. In the in vivo evaluation experiment, the scaffolds containing RBMSCs without prior induction were autologous implanted into the articular cartilage defects made by subchondral drilling. The repairing ability was evaluated. After 2 months, chondrocyte-like cells with lacuna structure and corresponding ECM were found in the repaired sites without apparent inflammation. After 24 weeks, we could easily find cartilage structure the same with normal cartilage in the repair site. In conclusion, it was shown that the scaffolds in combination of in vivo conditions can induce RBMSCs into chondrocytes in repaired area and would be a possible method for articular cartilage repair in clinic and cartilage tissue engineering.     

Compromised bone healing following spacer removal in a rat femoral defect model

Purpose: The clinical demand for bone grafting materials necessitated the development of animal models. Critical size defect model has been criticized recently, mainly for its inaccuracy. Our objective was to develop a dependable animal model that would provide compromised bone healing, and would allow the investigation of bone substitutes. Methods: In the first group a critical size defect was created in the femur of adult male Wistar rats, and a non-critical defect in the remaining animals (Groups II, III and IV). The defect was left empty in group II, while in groups III and IV a spacer was interposed into the gap. Osteoblast activity was evaluated by NanoSPECT/CT imaging system. New bone formation and assessment of a union or non-union was observed by μCT and histology. Results: The interposition model proved to be highly reproducible and provided a bone defect with compromised bone healing. Significant bone regeneration processes were observed four weeks after removal of the spacer. Conclusion: Our results have shown that when early bone healing is inhibited by the physical interposition of a spacer, the regeneration process is compromised for a further 4 weeks and results in a bone defect during the time-course of the study.     

Changes in collagen structure under the influence of mechanical dispersion

Air-dry collagen isolated from cattle retinal layer by means of alkaline-salt treatment was crushed in a laboratory vibro-mill at 80-150 degrees K. Mechanochemical transformations were studied by means of viscosimetry, polarimetry, ESR-spectroscopy and electron microscopy. Mechanical tensions induce breakage of covalent bonds of polypeptide chains, accompanied by a decrease of protein molecular mass, and of lateral interactions, which results in loosening of collagen structure and partial denaturation.