Proteoglycan and collagen contribution to the strain-rate-dependent mechanical behaviour of knee and shoulder cartilage.

The contribution of the proteoglycan to the strain-rate-dependent mechanical behaviour of cartilage tissues has been suggested to decrease with an increase in the strain-rate. On the other hand, the contribution from the collagen network has been suggested to increase as the strain-rate increases. These conclusions are drawn mainly based on numerical studies conducted on high-load-bearing knee cartilage tissues, while experimental evidence of these behaviours have not been demonstrated previously. Further, in contrast to the reported findings on high-load bearing knee cartilage, our previous study on the low-load-bearing kangaroo shoulder cartilage indicated that proteoglycan and collagen contribution remained steady as the strain-rate increases. Therefore, in the present study, we experimentally investigate the contribution of proteoglycan and collagen network to the strain-rate-dependent behaviour of the kangaroo knee cartilage, and plausible reasons for the differences observed in relation to the kangaroo shoulder cartilage. Firstly, in order to quantify the contribution of proteoglycans and collagen network, the indentation testings on normal, proteoglycan, and collagen-degraded kangaroo knee cartilage were conducted at different strain-rates. Then, structural and compositional differences between the kangaroo knee and shoulder cartilage were assessed qualitatively through polarised light microscopy (PLM) imaging and histological staining. Identified differences in the collagen architecture and proteoglycan composition were incorporated in a fibril-reinforced porohyperelastic Finite Element (FE) model with the objective of explaining the mechanisms underlying differences observed between the two tissues. Experimental results on knee cartilage indicated that when the strain-rate increases, proteoglycan contribution decreases while collagen contribution increases, where statistically significant differences were identified at each strain-rate (p < 0.05). PLM images revealed a sizable deep zone in the kangaroo knee cartilage where collagen fibrils were oriented perpendicular to the subchondral bone. On the other hand, no such apparent deep zone was observed in the shoulder cartilage. FE model confirmed that the biomechanical differences observed in the knee and shoulder cartilage are due to the differences in the collagen fibril arrangement in the deep zone. From these results, it can be concluded that in high-load-bearing cartilage tissues, the collagen network in the deep zone assists in increasing the stiffness of tissue with strain-rate and plays a significant role in supporting transient loads. This, in turn, helps protect the solid matrix against large distortions and strains at the subchondral junction, pointing to the importance of the collagen network in deep zone in assisting high-load-bearing cartilage tissues.

Impact of Nanoparticle Uptake on the Biophysical Properties of Cell for Biomedical Engineering Applications.

Nanomaterials are currently the state-of-the-art in the development of advanced biomedical devices and applications where classical approaches have failed. To date, majority of the literature on nanomaterial interaction with cells have largely focused on the biological responses of cells obtained via assays, with little interest on their biophysical responses. However, recent studies have shown that the biophysical responses of cells, such as stiffness and adhesive properties, play a significant role in their physiological function. In this paper, we investigate cell biophysical responses after uptake of nanoparticles. Atomic force microscopy was used to study changes in cell stiffness and adhesion upon boron nitride (BN) and hydroxyapatite (HAP) nanoparticle uptake. Results show increase in cell stiffness with varying nanoparticle (BN and HAP) concentration, while a decrease in cell adhesion trigger by uptake of HAP. In addition, changes in the biochemical response of the cell membrane were observed via Raman spectroscopy of nanoparticle treated cells. These findings have significant implications in biomedical applications of nanoparticles, e.g. in drug delivery, advanced prosthesis and surgical implants.

Potential enhancement of articular cartilage histological grading with collagen integrity.

BACKGROUND: Histological evaluation of articular cartilage, such as using the Mankin scoring system, is the gold standard for characterization of tissue integrity. This scoring system takes into account several parameters indicative of the tissue's health; however, the collagen integrity, which is a primary indicator of cartilage health is not taken into consideration. Thus, there is need to enhance histological grading of articular cartilage by incorporating explicit scoring of collagen degeneration into the Modified Mankin grading system. This paper explores a new histological grading parameter for collagen network degradation and how this information can be used to augment a widely used grading scheme like the Modified Mankin grading system. METHODS: Intact and degenerated human cartilage were examined histologically and then subjected to second harmonic generation imaging, leading to qualitative and quantitative description of collagen disruption emanating from the surface to subsurface layers of the tissue. This data was then incorporated into the Modified Mankin grading system. FINDINGS: Second harmonic generation image analysis reveals a relationship between changes in collagen architecture and histologically observed tissue disruption in degenerated articular cartilage. INTERPRETATION: Histological tissue disruption in degenerated human articular cartilage is directly related to the reorganization of collagen fibrils in the form of intense fibril aggregation, either as a result of degeneration or aging. This method of mapping disrupted tissue regions to quantitative collagen fibril damage can be coded into cartilage grading systems and could inform clinical practice and scientific research.

Characterization of Articular Cartilage Recovery and Its Correlation with Optical Response in the Near-Infrared Spectral Range.

OBJECTIVES: In this study, we examine the capacity of a new parameter, based on the recovery response of articular cartilage, to distinguish between healthy and damaged tissues. We also investigate whether or not this new parameter correlates with the near-infrared (NIR) optical response of articular cartilage. DESIGN: Normal and artificially degenerated (proteoglycan-depleted) bovine cartilage samples were nondestructively probed using NIR spectroscopy. Subsequently they were subjected to a load and unloading protocol, and the recovery response was logged during unloading. The recovery parameter, elastic rebound ( ER), is based on the strain energy released as the samples underwent instantaneous elastic recovery. RESULTS: Our results reveal positive relationship between the rebound parameter and cartilage proteoglycan content (normal samples: 2.20 ± 0.10 N mm; proteoglycan-depleted samples: 0.50 ± 0.04 N mm for 1 hour of enzymatic treatment and 0.13 ± 0.02 N mm for 4 hours of enzymatic treatment). In addition, multivariate analysis using partial least squares regression was employed to investigate the relationship between ER and NIR spectral data. The results reveal significantly high correlation ( R2cal = 98.35% and R2val = 79.87%; P < 0.0001), with relatively low error (14%), between the recovery and optical response of cartilage in the combined NIR regions 5,450 to 6,100 cm-1 and 7,500 to 12,500 cm-1. CONCLUSION: We conclude that ER can indicate the mechanical condition and state of health of articular cartilage. The correlation of ER with cartilage optical response in the NIR range could facilitate real-time evaluation of the tissue's integrity during arthroscopic surgery and could also provide an important tool for cartilage assessment in tissue engineering and regeneration research.

Bactericidal Effects of Natural Nanotopography of Dragonfly Wing on Escherichia coli.

Nanotextured surfaces (NTSs) are critical to organisms as self-adaptation and survival tools. These NTSs have been actively mimicked in the process of developing bactericidal surfaces for diverse biomedical and hygiene applications. To design and fabricate bactericidal topographies effectively for various applications, understanding the bactericidal mechanism of NTS in nature is essential. The current mechanistic explanations on natural bactericidal activity of nanopillars have not utilized recent advances in microscopy to study the natural interaction. This research reveals the natural bactericidal interaction between E. coli and a dragonfly wing's (Orthetrum villosovittatum) NTS using advanced microscopy techniques and proposes a model. Contrary to the existing mechanistic models, this experimental approach demonstrated that the NTS of Orthetrum villosovittatum dragonfly wings has two prominent nanopillar populations and the resolved interface shows membrane damage occurred without direct contact of the bacterial cell membrane with the nanopillars. We propose that the bacterial membrane damage is initiated by a combination of strong adhesion between nanopillars and bacterium EPS layer as well as shear force when immobilized bacterium attempts to move on the NTS. These findings could help guide the design of novel biomimetic nanomaterials by maximizing the synergies between biochemical and mechanical bactericidal effects.

Preparation of bone-implants by coating hydroxyapatite nanoparticles on self-formed titanium dioxide thin-layers on titanium metal surfaces.

Preparation of hydroxyapatite coated custom-made metallic bone-implants is very important for the replacement of injured bones of the body. Furthermore, these bone-implants are more stable under the corrosive environment of the body and biocompatible than bone-implants made up of pure metals and metal alloys. Herein, we describe a novel, simple and low-cost technique to prepare biocompatible hydroxyapatite coated titanium metal (TiM) implants through growth of self-formed TiO2 thin-layer (SFTL) on TiM via a heat treatment process. SFTL acts as a surface binder of HA nanoparticles in order to produce HA coated implants. Colloidal HA nanorods prepared by a novel surfactant-assisted synthesis method, have been coated on SFTL via atomized spray pyrolysis (ASP) technique. The corrosion behavior of the bare and surface-modified TiM (SMTiM) in a simulated body fluid (SBF) medium is also studied. The highest corrosion rate is found to be for the bare TiM plate, but the corrosion rate has been reduced with the heat-treatment of TiM due to the formation of SFTL. The lowest corrosion rate is recorded for the implant prepared by heat treatment of TiM at 700 °C. The HA-coating further assists in the passivation of the TiM in the SBF medium. Both SMTiM and HA coated SMTiM are noncytotoxic against osteoblast-like (HOS) cells and are in high-bioactivity. The overall production process of bone-implant described in this paper is in high economic value.

Investigation of the Effects of Extracellular Osmotic Pressure on Morphology and Mechanical Properties of Individual Chondrocyte.

It has been demonstrated that most cells of the body respond to osmotic pressure in a systematic manner. The disruption of the collagen network in the early stages of osteoarthritis causes an increase in water content of cartilage which leads to a reduction of pericellular osmolality in chondrocytes distributed within the extracellular environment. It is therefore arguable that an insight into the mechanical properties of chondrocytes under varying osmotic pressure would provide a better understanding of chondrocyte mechanotransduction and potentially contribute to knowledge on cartilage degeneration. In this present study, the chondrocyte cells were exposed to solutions with different osmolality. Changes in their dimensions and mechanical properties were measured over time. Atomic force microscopy (AFM) was used to apply load at various strain-rates and the force-time curves were logged. The thin-layer elastic model was used to extract the elastic stiffness of chondrocytes at different strain-rates and at different solution osmolality. In addition, the porohyperelastic (PHE) model was used to investigate the strain-rate-dependent responses under the loading and osmotic pressure conditions. The results revealed that the hypo-osmotic external environment increased chondrocyte dimensions and reduced Young's modulus of the cells at all strain-rates tested. In contrast, the hyper-osmotic external environment reduced dimensions and increased Young's modulus. Moreover, using the PHE model coupled with inverse FEA simulation, we established that the hydraulic permeability of chondrocytes increased with decreasing extracellular osmolality which is consistent with previous work in the literature. This could be due to a higher intracellular fluid volume fraction with lower osmolality.

Microscale consolidation analysis of relaxation behavior of single living chondrocytes subjected to varying strain-rates.

Besides the elastic stiffness, the relaxation behavior of single living cells is also of interest of various researchers when studying cell mechanics. It is hypothesized that the relaxation response of the cells is governed by both intrinsic viscoelasticity of the solid phase and fluid-solid interactions mechanisms. There are a number of mechanical models have been developed to investigate the relaxation behavior of single cells. However, there is lack of model enable to accurately capture both of the mechanisms. Therefore, in this study, the porohyperelastic (PHE) model, which is an extension of the consolidation theory, combined with inverse Finite Element Analysis (FEA) technique was used at the first time to investigate the relaxation response of living chondrocytes. This model was also utilized to study the dependence of relaxation behavior of the cells on strain-rates. The stress-relaxation experiments under the various strain-rates were conducted with the Atomic Force Microscopy (AFM). The results have demonstrated that the PHE model could effectively capture the stress-relaxation behavior of the living chondrocytes, especially at intermediate to high strain-rates. Although this model gave some errors at lower strain-rates, its performance was acceptable. Therefore, the PHE model is properly a promising model for single cell mechanics studies. Moreover, it has been found that the hydraulic permeability of living chondrocytes reduced with decreasing of strain-rates. It might be due to the intracellular fluid volume fraction and the fluid pore pressure gradients of chondrocytes were higher when higher strain-rates applied.

Spatial mapping of proteoglycan content in articular cartilage using near-infrared (NIR) spectroscopy.

Diagnosis of articular cartilage pathology in the early disease stages using current clinical diagnostic imaging modalities is challenging, particularly because there is often no visible change in the tissue surface and matrix content, such as proteoglycans (PG). In this study, we propose the use of near infrared (NIR) spectroscopy to spatially map PG content in articular cartilage. The relationship between NIR spectra and reference data (PG content) obtained from histology of normal and artificially induced PG-depleted cartilage samples was investigated using principal component (PC) and partial least squares (PLS) regression analyses. Significant correlation was obtained between both data (R(2) = 91.40%, p<0.0001). The resulting correlation was used to predict PG content from spectra acquired from whole joint sample, this was then employed to spatially map this component of cartilage across the intact sample. We conclude that NIR spectroscopy is a feasible tool for evaluating cartilage contents and mapping their distribution across mammalian joint.

Sterilizing tissue-materials using pulsed power plasma.

This paper investigates the potential of pulsed power to sterilize hard and soft tissues and its impact on their physico-mechanical properties. It hypothesizes that pulsed plasma can sterilize both vascular and avascular tissues and the transitive layers in between without deleterious effects on their functional characteristics. Cartilage/bone laminate was chosen as a model to demonstrate the concept, treated at low temperature, at atmospheric pressure, in short durations and in buffered environment using a purposed-built pulsed power unit. Input voltage and time of exposure were assigned as controlling parameters in a full factorial design of experiment to determine physical and mechanical alteration pre- and post-treatment. The results demonstrated that, discharges of 11 kV sterilized samples in 45 s, reducing intrinsic elastic modules from 1.4 ± 0.9 to 0.9 ± 0.6 MPa. There was a decrease of 14.1 % in stiffness and 27.8 % in elastic-strain energy for the top quartile. Mechanical impairment was directly proportional to input voltage (P value < 0.05). Bacterial inactivation was proportional to treatment time for input voltages above 32 V (P < 0.001; R Sq = 0.98). Thermal analysis revealed that helix-coil transition decelerated with exposure time and collagen fibrils were destabilized as denaturation enthalpy reduced by 200 µV. We concluded by presenting a safe operating threshold for pulsed power plasma as a feasible protocol for effective sterilization of connective tissues with varying level of loss in mechanical robustness which we argue to be acceptable in certain medical and tissue engineering application.

Cellular senescence and longevity of osteophyte-derived mesenchymal stem cells compared to patient-matched bone marrow stromal cells.

This study aimed to determine the cellular aging of osteophyte-derived mesenchymal cells (oMSCs) in comparison to patient-matched bone marrow stromal cells (bMSCs). Extensive expansion of the cell cultures was performed and early and late passage cells (passages 4 and 9, respectively) were used to study signs of cellular aging, telomere length, telomerase activity, and cell-cycle-related gene expression. Our results showed that cellular aging was more prominent in bMSCs than in oMSCs, and that oMSCs had longer telomere length in late passages compared with bMSCs, although there was no significant difference in telomere lengths in the early passages in either cell type. Telomerase activity was detectable only in early passage oMSCs and not in bMSCs. In osteophyte tissues telomerase-positive cells were found to be located perivascularly and were Stro-1 positive. Fifteen cell-cycle regulator genes were investigated and only three genes (APC, CCND2, and BMP2) were differentially expressed between bMSC and oMSC. Our results indicate that oMSCs retain a level of telomerase activity in vitro, which may account for the relatively greater longevity of these cells, compared with bMSCs, by preventing replicative senescence.

Characterization of a mesenchymal-like stem cell population from osteophyte tissue.

Osteophytes are a distinct feature of osteoarthritis (OA). Their formation may be related to pluripotential cells in the periosteum responding to stimulus during OA. This study aimed to isolate stem cells from osteophyte tissues and to characterize their phenotype, proliferation, and differentiation potential, as well as their immunomodulatory properties. Osteophyte-derived cells were isolated from osteophyte tissue samples collected during knee replacement surgery. These cells were characterized by the expression of cell-surface antigens, differentiation potential into mesenchymal lineages, growth kinetics, and modulation of alloimmune responses. Multipotential stem cells were identified from all osteophyte samples, namely osteophyte-derived mesenchymal stem cells (oMSCs). The surface antigen expression of oMSCs was consistent with that of MSCs; they lacked the hematopoietic and common leukocyte markers (CD34, CD45) while expressing those related to adhesion (CD29, CD166, CD44) and stem cells (CD90, CD105, CD73). The proliferation capacity of oMSCs in culture was superior to that of bone marrow-derived MSCs (bMSCs), and these cells readily differentiated into tissues of the mesenchymal lineages. oMSCs also demonstrated the ability to suppress allogeneic T cell proliferation, which was associated with the expression of the tryptophan-degrading enzyme indoleamine 2,3-dioxygenase (IDO). Our results showed that osteophyte-derived cells had similar properties to MSCs in the expression of antigen phenotype, differential potential, and suppression of alloimmune response. Furthermore, when compared to bMSCs, oMSCs maintained a higher proliferative capacity, which may offer new insights of the tissue formation and potentially an alternative source for therapeutic stem cell-based tissue regeneration.