Biorheology - Volume 43, issue 3-4

Authors: Kopakkala-Tani, Milla | Leskinen, Jarkko J. | Karjalainen, Hannu M. | Karjalainen, Tero | Hynynen, Kullervo | Töyräs, Juha | Jurvelin, Jukka S. | Lammi, Mikko J.

Article Type: Research Article

Abstract: Mechanical forces can stimulate the production of extracellular matrix molecules. We tested the efficacy of ultrasound to increase proteoglycan synthesis in bovine primary chondrocytes. The ultrasound-induced temperature rise was measured and its contribution to the synthesis was investigated using bare heat stimulus. Chondrocytes from five cellular isolations were exposed in triplicate to ultrasound (1 MHz, duty cycle 20%, pulse repetition frequency 1 kHz) at average intensity of 580 mW/cm2 for 10 minutes daily for 1–5 days. Temperature evolution was recorded during the sonication and corresponding temperature history was created using a controllable water bath. This exposure profile was used …in 10-minute-long heat treatments of chondrocytes. Heat shock protein 70 (Hsp70) levels after one-time treatment to ultrasound and heat was analyzed by Western blotting, and proteoglycan synthesis was evaluated by 35 S-sulfate incorporation. Ultrasound treatment did not induce Hsp70, while heat treatment caused a slight heat stress response. Proteoglycan synthesis was increased approximately 2-fold after 3–4 daily ultrasound stimulations, and remained at that level until day 5 in responsive cell isolates. However, chondrocytes from one donor cell isolation out of five remained non-responsive. Heat treatment alone did not increase proteoglycan synthesis. In conclusion, our study confirms that pulsed ultrasound stimulation can induce proteoglycan synthesis in chondrocytes. Show more

Keywords: Ultrasound treatment, heat, proteoglycans, chondrocyte

Citation: Biorheology, vol. 43, no. 3-4, pp. 271-282, 2006

Authors: Sivan, S. | Neidlinger-Wilke, C. | Würtz, K. | Maroudas, A. | Urban, J.P.G.

Article Type: Research Article

Abstract: The intervertebral discs are large cartilaginous structures situated between the vertebral bodies, occupying around one third of the length of the spinal column. They act as the joints of the spine and carry mechanical load arising from body weight and muscle activity. Loads change with every alteration of posture and activity and the discs thus undergo a diurnal loading pattern with high loads on the discs during the day's activity and low loads on it at night during rest. As the disc is an osmotic system, around 25% of the disc's fluid is expressed and re-imbibed during each diurnal cycle …with consequent changes in the osmotic environment of the disc cells. Here, present information on the effect of osmotic changes in disc cell metabolism is reviewed; results indicate that prevailing osmolarity is a powerful regulator of disc cell activity. Show more

Citation: Biorheology, vol. 43, no. 3-4, pp. 283-291, 2006

Authors: Marques, Mara Rúbia | Hajjar, Denise | Oliveira Crema, Virgínia | Kimura, Edna Teruko | Santos, Marinilce Fagundes

Article Type: Research Article

Abstract: We have previously shown that a mandibular propulsive appliance (MPA) stimulates cell proliferation and the synthesis of growth factors in the rat condylar cartilage. The aim of this study was to evaluate the effects of a MPA in the distribution of the integrin subunits α1 and α2 in this cartilage. Twenty eight days-old male Wistar rats were divided into treated (T) and age-matched control groups (C). Treated rats wore the appliance during 3, 5, 7, 9, 11, 15, 20, 30 and 35 days. The condyles were fixed, decalcified and paraffin-embedded. The distribution of α1 and α2 was studied by immunohistochemistry. …Alpha1 distribution was uniform along the cartilage, increasing in 48 days-old rats (C20). Treated animals anticipated this increase to the age of 36 days (T9). The number of α2-positive cells was increased in C9 in the anterior condylar region, in C9 and C20 in the middle region and showed no differences in the posterior region. The MPA apparently abolished all variations, leading to a single increase at T30 in all regions. These results suggest that integrins containing the α1 and α2 subunits are modulated by forces promoted by the MPA, participating of the biological response to this therapy. Show more

Keywords: Functional orthopedics, condyle, alpha1, alpha2, integrin

Citation: Biorheology, vol. 43, no. 3-4, pp. 293-302, 2006

Authors: Benallaoua, Mourad | Richette, Pascal | François, Mathias | Tsagris, Lydia | Revel, Michel | Corvol, Maité | Poiraudeau, Serge | Savouret, Jean-François | Rannou, François

Article Type: Research Article

Abstract: Proteoglycan production is one of the major extracellular matrix components implicated in the dynamic process of intervertebral disc degeneration. Mechanical stress is an important modulator of the degeneration, but the underlying molecular mechanism at the proteoglycan level remains unclear. The aim of this work was to study the regulation of proteoglycan production by cyclic tensile stretch applied to intervertebral disc annulus fibrosus cells. Matrix metalloproteinases do not seem to be implicated in the regulation of proteoglycan production. By contrast, nitrite oxide production is induced by cyclic tensile stretch, in a time, intensity, and frequency dependant manner. Using a non-specific nitric …oxide synthases inhibitor [NG -methyl-L-arginine (L-NMA)], we suppress totally the inhibition of proteoglycan production induced by cyclic tensile stretch suggesting the implication of nitric oxide synthases in the observed phenomenon. Introducing the transcriptional inhibitor 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole or a more specific inhibitor of nitric oxide synthases II [N-iminoethyl-L-lysine (L-NIL)] did not affect the decreased proteoglycan production, which suggests a post-translational regulation. In contrast, N-omega nitro-L-arginine (L-NNA) a more specific inhibitor of NOS I and III abrogated the cyclic tensile stretch-dependant inhibition of proteoglycan production. These results suggest that cyclic tensile stretch regulates proteoglycan production through a post-translational mechanism involving nitrite oxide. This result could be of interest in the development of local therapeutic strategies aimed at controlling intervertebral disc degeneration. Show more

Keywords: Intervertebral disc, mechanical stretch, annulus fibrosus, proteoglycan, chondrocyte, nitrite oxide

Citation: Biorheology, vol. 43, no. 3-4, pp. 303-310, 2006

Authors: Hajjar, Denise | Santos, Marinilce F. | Kimura, Edna Teruko

Article Type: Research Article

Abstract: Functional orthopedic appliances correct dental malocclusion partially by exerting indirect mechanical stimulus on the condylar cartilage, modulating growth and the adaptation of orofacial structures. However, the exact nature of the biological responses to this therapy is not well understood. Insulin-like growth factors I and II (IGF-I and II) are important local factors during growth and differentiation in the condylar cartilage [D. Hajjar, M.F. Santos and E.T. Kimura, Propulsive appliance stimulates the synthesis of insulin-like growth factors I and II in the mandibular condylar cartilage of young rats, Arch. Oral Biol. 48 (2003), 635–642]. The bioefficacy of IGFs at the cellular …level is modulated by IGF binding proteins (IGFBP). The aim of this study was to verify the mRNA and protein expression of IGFBP-3, IGFBP-4, IGFBP-5 and IGFBP-6 in the condylar cartilage of young male Wistar rats that used a mandibular propulsive appliance for 3, 9, 15, 20, 30 or 35 days. For this purpose, sagittal sections of decalcified and paraffin-embedded condyles were submitted to immunohistochemistry and the condylar cartilage to RT–PCR. The control group showed a gradual increase in the protein expression of all IGFBPs, except IGFBP-4. Following use of the appliance, IGFBP-3 and IGFBP-6 expression decreased in the early stage of the treatment. At 20 days of treatment there was a decline in the IGFs and IGFBP-3, IGFBP-4 and IGFBP-5 expression and at 30 days there was a peak in the IGFs and all IGFBPs expression except for IGFBP-3 where the peak was observed in the control animals. The expression patterns of all IGFBPs in the condylar cartilage were similar. The modulation of IGFBP-3, -4, -5 and -6 expression in the condylar cartilage in response to the propulsive appliance suggests that those peptides are involved in the mandibular adaptation during this therapy. Show more

Keywords: Insulin-like growth factor binding protein, mechanical stress, mandibular condyle, cartilage, functional orthopedics

Citation: Biorheology, vol. 43, no. 3-4, pp. 311-321, 2006

Authors: Yao, Hai | Gu, Wei Yong

Article Type: Research Article

Abstract: A 3D finite element model for charged hydrated soft tissues containing charged/uncharged solutes was developed based on the multi-phasic mechano-electrochemical mixture theory (Lai et al., J. Biomech. Eng. 113 (1991), 245–258; Gu et al., J. Biomech. Eng. 120 (1998), 169–180). This model was applied to analyze the mechanical, chemical and electrical signals within the human intervertebral disc during an unconfined compressive stress relaxation test. The effects of tissue composition [e.g., water content and fixed charge density (FCD)] on the physical signals and the transport rate of fluid, ions and nutrients were investigated. The numerical simulation showed that, during disc compression, …the fluid pressurization was more pronounced at the center (nucleus) region of the disc while the effective (von Mises) stress was higher at the outer (annulus) region. Parametric analyses revealed that the decrease in initial tissue water content (0.7–0.8) increased the peak stress and relaxation time due to the reduction of permeability, causing greater fluid pressurization effect. The electrical signals within the disc were more sensitive to FCD than tissue porosity, and mechanical loading affected the large solute (e.g., growth factor) transport significantly, but not for small solute (e.g., glucose). Moreover, this study confirmed that the interstitial fluid pressurization plays an important role in the load support mechanism of IVD by sharing more than 40% of the total load during disc compression. This study is important for understanding disc biomechanics, disc nutrition and disc mechanobiology. Show more

Keywords: Intervertebral disc, triphasic theory, finite element method, solute transport, soft tissue mechanics

Citation: Biorheology, vol. 43, no. 3-4, pp. 323-335, 2006

Authors: Chen, M.-Y. | Jeng, L. | Sun, Y.-L. | Zhao, C.-F. | Zobitz, M.E. | Moran, S.L. | Amadio, P.C. | An, K.-N.

Article Type: Research Article

Abstract: Knowledge of the adaptation of the soft tissue to mechanical factors and biomolecules would be essential to better understand the mechanism of tendon injury and to improve the outcome of tendon repair. The responses to these factors could be different for the distinct types of cells in the tendon: cells from the tendon sheath, fibroblasts from the epitenon surface, or fibroblasts from the internal endotenon. In this study, we examined the mechanical and histological characteristics of the rate of contraction of the collagen gel seeded with epitenon and endotenon fibroblasts. The rate of contraction and the mechanical property of the …contracted construct depend on the gel concentration and also the treatment of TGF-β1 . Show more

Keywords: Gel concentration, TGF-β, fibroblast, tendon, cell culture, biomechanics

Citation: Biorheology, vol. 43, no. 3-4, pp. 337-345, 2006

Authors: Kato, Yukio | Nakamura, Shigeo | Nishimura, Masahiro

Article Type: Research Article

Abstract: Hyaluronan (HA) has viscoelastic, anti-inflammatory and protective actions in joint tissues, and is being widely used for treatment of OA and RA patients. However, the mechanisms underlying the pharmacological action of HA on OA and RA have not been fully understood. In this article, we review the molecular weight-dependent, anti-inflammatory actions of HA preparations – produced in Japan – in joint tissues, and show that the molecular weight of HA, but not its concentration, is crucial for maintenance of cartilage elasticity.

Keywords: Hyaluronan, aggrecan, cartilage, OA, RA, elasticity

Citation: Biorheology, vol. 43, no. 3-4, pp. 347-354, 2006

Authors: Archer, Charles W. | Buxton, Paul | Hall, Brian K. | Francis-West, Philippa

Article Type: Research Article

Abstract: This article has been retracted. The original article can be found on: http://dev.biologists.org/cgi/reprint/130/19/4729.

Citation: Biorheology, vol. 43, no. 3-4, pp. 355-370, 2006

Authors: Huselstein, C. | de Isla, N. | Kolopp-Sarda, M.N. | Kerdjoudj, H. | Muller, S. | Stoltz, J.F.

Article Type: Research Article

Abstract: The cartilage is a hydrated connective tissue in joints that withstands and distributes mechanical forces. The chondrocytes utilize mechanical signals to regulate their metabolic activity through complex biological and biophysical interactions with the extracellular matrix (ECM). The aim of this work was to study the influence of mechanical stress on cells behavior cultured in 3D biosystems (alginate and alginate supplemented with hyaluronate). After mechanical stimulation, cell viability and cell death process were the main studied parameters. Our results indicated that viability and cell cycle progression were inhibited under mechanical stimulation, as far as the extracellular matrix was not yet synthesized. …In contrast, on day 21, the mechanical stimulation had positive effect on these parameters. Show more

Keywords: Chondrocyte, alginate hydrogel, cell viability, mechanical stimulation, extracellular matrix

Citation: Biorheology, vol. 43, no. 3-4, pp. 371-375, 2006

Authors: Botter, S.M. | van Osch, G.J.V.M. | Waarsing, J.H. | Day, J.S. | Verhaar, J.A.N. | Pols, H.A.P. | van Leeuwen, J.P.T.M. | Weinans, H.

Article Type: Research Article

Abstract: In the past few years there has been a considerable interest in the role of bone in osteoarthritis. Despite the increasing evidence of the involvement of bone in osteoarthritis, it remains very difficult to attribute the cause or effect of changes in subchondral bone to the process of osteoarthritis. Although osteoarthritis in mice provides a useful model to study changes in the subchondral bone, detailed quantification of these changes is lacking. Therefore, the goal of this study was to quantify subchondral bone changes in a murine osteoarthritis model by use of micro-computed tomography (micro-CT). We induced osteoarthritis-like characteristics in the …knee joints of mice using collagenase injections, and after four weeks we calculated various 3D morphometric parameters in the epiphysis of the proximal tibia. The collagenase injections caused cartilage damage, visible in histological sections, particularly on the medial tibial plateau. Micro-CT analysis revealed that the thickness of the subchondral bone plate was decreased both at the lateral and the medial side. The trabecular compartment demonstrated a small but significant reduction in bone volume fraction compared to the contralateral control joints. Trabeculae in the collagenase-injected joints were thinner but their shape remained rod-like. Furthermore, the connectivity between trabeculae was reduced and the trabecular spacing was increased. In conclusion, four weeks after induction of osteoarthritis in the murine knee subtle but significant changes in subchondral bone architecture could be detected and quantified in 3D with micro-CT analysis. Show more

Citation: Biorheology, vol. 43, no. 3-4, pp. 379-388, 2006

Authors: Quasnichka, Helen L. | Anderson-MacKenzie, Janet M. | Bailey, Allen J.

Article Type: Research Article

Abstract: There is increasing recognition that osteoarthritis (OA) is a complex disease involving the whole synovial joint, rather than the articular cartilage alone, however its aetiology and pathogenesis is not understood. Our initial studies revealed elevated turnover of bone and ligament collagen in human and mouse OA, respectively. To investigate the relative appearance of pathology in cartilage, bone and ligament, we studied the progression of spontaneous OA in the Dunkin–Hartley (DH) guinea pig knee, and compared with age-matched control Bristol Strain 2 (BS2) knees. The classical radiographic OA score of the DH knees compared to BS2 knees was 2-fold higher at …24 weeks of age. The patella perimeter and subchondral bone density was significantly greater in the DHs at 24 and 36 weeks compared to BS2. The femoral intercondylar notch width was found to be significantly lower in the DHs at 24 and 36 weeks, compared to BS2, indicating bone remodelling at the cruciate ligament (CL) insertion site. We found significantly greater laxity of the DH anterior CL at 12, 16 and 20 weeks compared to BS2. This elevated laxity was associated with increased remodelling of the CLs, based on markers of collagen turnover, and occurred prior to bone and cartilage pathology. We propose that the laxity of the CL leads to remodelling of the subchondral bone, and intercondylar notch, due to a change in load through the joint. Remodelling of the CLs and bone occurs prior to and concomitant with histopathological changes in the articular cartilage respectively, demonstrating the fundamental role of the ligament and subchondral bone in the aetiology of knee OA. Show more

Keywords: Osteoarthritis, subchondral bone, ligaments, collagen, metabolism, biomechanics, notch width

Citation: Biorheology, vol. 43, no. 3-4, pp. 389-397, 2006

Article Type: Other

Citation: Biorheology, vol. 43, no. 3-4, pp. 401-402, 2006

Authors: Watrin-Pinzano, Astrid | Etienne, Stéphanie | Grossin, Laurent | Gaborit, Nadège | Cournil-Henrionnet, Christel | Mainard, Didier | Netter, Patrick | Gillet, Pierre | Galois, Laurent

Article Type: Research Article

Keywords: Cartilage, osteoarthritis, apoptosis, caspase, experimental model

Citation: Biorheology, vol. 43, no. 3-4, pp. 403-412, 2006

Authors: Chowdhury, Tina T. | Bader, Dan L. | Lee, David A.

Article Type: Research Article

Abstract: . NO and PGE2 are inflammatory mediators derived from the inducible iNOS and COX enzymes and are potentially important pharmacological targets in OA. Both mechanical loading and IL-1β will influence the release of . NO and PGE2 . Accordingly, the current study examines the effect of dynamic compression on . NO and PGE2 release by human chondrocytes cultured in agarose constructs in the presence and absence of selective iNOS and COX-2 inhibitors. The current data demonstrate that IL-1β induced nitrite and PGE2 release and inhibited [3 H]-thymidine and 35 SO4 incorporation. Inhibitor experiments indicate that 1400W …and NS-398 either partially reversed or abolished IL-1β induced nitrite and PGE2 release. IL-1β induced inhibition of cell proliferation and proteoglycan synthesis was partially reversed with 1400W but was not influenced by NS-398. For the dynamic loading experiments, 1400W and NS-398 either reduced or abolished the compression-induced inhibition of . NO and PGE2 release in the presence of IL-1β. The IL-1β induced inhibition of cell proliferation was not influenced by 1400W or NS-398 whereas strain-induced stimulation of proteoglycan synthesis in the presence of IL-1β was enhanced by 1400W. The data obtained using human chondrocytes demonstrate that IL-1β induced . NO and PGE2 release via an iNOS-driven-COX-2 inter-dependent pathway. This response could be reversed by dynamic compression. These data indicate interactions exist between the NOS and COX pathways, a finding which will provide new insights in the development of pharmacological or biophysical treatments for cartilage disorders such as OA. Show more

Keywords: Nitric oxide, PGE$_{2}$, mechanotransduction, osteoarthritis

Citation: Biorheology, vol. 43, no. 3-4, pp. 413-429, 2006

Authors: Schumann, D. | Kujat, R. | Zellner, J. | Angele, M.K. | Nerlich, M. | Mayr, E. | Angele, P.

Article Type: Research Article

Abstract: This study examined the effects of low intensity pulsed ultrasound (LIPUS) on human bone marrow-derived mesenchymal stem cells undergoing chondrogenic differentiation. Aggregates of mesenchymal stem cells and mesenchymal stem cells seeded in three dimensional matrices were cultured in a defined chondrogenic medium and subjected to LIPUS for the first 7 days of culture. At 1, 7, 14 and 21 days, samples were harvested for histology, immunohistochemistry, RT-PCR, and quantitative DNA and matrix macromolecule analysis. Cell aggregates with daily treatment for 20 minutes showed no significant differences for proteoglycan and collagen content during chondrogenic differentiation. However ultrasound application for 40 minutes …daily resulted in a statistically significant increase of the proteoglycan and collagen content after 21 days in culture. Aggregates treated for 20 minutes daily showed decreased expression of chondrogenic genes at all time points. In contrast, 40 minutes of daily treatment of aggregates resulted in a significant increase of chondrogenic marker genes after an initial decrease at day 7 with time in culture. Ultrasound treated cell-scaffold constructs showed a significant increase of chondrogenic marker gene expression and extracellular matrix deposition. This study indicates that LIPUS can be used to enhance the chondrogenesis of mesenchymal stem cells in cell aggregates and cell-scaffold constructs. We have found a dependency on the specific treatment parameters. We hypothesize that LIPUS can be used for an improved in vitro preparation of optimized tissue engineering implants for cartilage repair. Furthermore this non-invasive method could also be of potential use in vivo for regenerative therapy of cartilage in the future. Show more

Keywords: Chondrogenesis, stem cell, bone marrow, mechanobiology, ultrasound

Citation: Biorheology, vol. 43, no. 3-4, pp. 431-443, 2006

Authors: Hiraoka, Koji | Grogan, Shawn | Olee, Tsaiwei | Lotz, Martin

Article Type: Research Article

Abstract: The transmembrane receptor Notch-1 regulates cell fate and differentiation and was suggested to identify a cell type with progenitor characteristics in newborn bovine articular cartilage. We show that Notch-1 is expressed on >70% of BM-MSC in early passage monolayer culture. We also demonstrate that normal articular cartilage contains Notch-1+ cells and that the frequency is increased in OA. Most Notch-1+ cells in OA cartilage are located in the clusters of proliferating cells. These findings indicate that multipotential mesenchymal progenitor cells are present in articular cartilage from adult humans and that their frequency is increased in OA. This observation has implications …for understanding the intrinsic repair capacity of articular cartilage and raises the possibility that these progenitor cells might be involved in the pathogenesis of arthritis. Show more

Citation: Biorheology, vol. 43, no. 3-4, pp. 447-454, 2006

Authors: Campbell, Jonathan J. | Lee, David A. | Bader, Dan L.

Article Type: Research Article

Abstract: This study tests the hypothesis that dynamic compressive strain selectively enhances chondrogenic differentiation by human mesenchymal stem cells (MSCs). Primary MSCs were isolated and expended in monolayer culture. The cells were seeded in alginate constructs or in pellet culture. The time course of chondrogenic differentiation was assessed by real-time QPCR of mRNA expression analysis for cartilage specific markers. Collagen types II and X mRNA, not present in undifferentiated MSCs, were detectable by 2–4 days of chondrogenic induction and continued to rise significantly throughout the culture period of 10 days (p<0.001). Basal levels of gene expression for Sox-9 and aggrecan were …evident in undifferentiated MSCs, although chondrogenic induction for a period of 8 days resulted in an increased trend in the gene expression levels. The alginate system was also used in mechanical conditioning studies. Dynamic compression was applied, in an intermittent regimen, at a strain amplitude of 15% and frequency of 1 Hz in the presence and absence of 10 ng/ml TGFβ3 , for a period of 8 days. Results indicated significant changes in the levels of mRNA expression for the chondrogenic markers. For example, by day 8, the application of the strain regimen alone caused an up-regulation in all the chondrogenic markers compared to the control samples (no TGFβ, no compression). However, the combined effects of strain and TGFβ on these markers were more complex than purely additive. Show more

Citation: Biorheology, vol. 43, no. 3-4, pp. 455-470, 2006

Authors: Facchini, Andrea | Lisignoli, Gina | Cristino, Sandra | Roseti, Livia | De Franceschi, Luciana | Marconi, Emanuele | Grigolo, Brunella

Article Type: Research Article

Abstract: The development of improved methods for treatment of chondral defects using autologous cells in combination with biomaterials leads to a new generation of implantable devices. Their association gives rise to a hybrid construct combining biological and material components that can be specifically committed. The comprehension of cellular and molecular mechanisms of cartilage repair and the use of biomaterials in combination with chondrocytes or mesenchymal stem cells in the treatment of cartilage defects has opened a new era of therapeutical strategies. Recently, their applicability in the treatment of early lesions in osteoarthritis is under investigation. To obtain new information on the …behaviour of chondrocytes and mesenchymal stem cells grown on a hyaluronan derivative scaffold (Hyaff® -11) already used in cartilage repair, we analysed a series of molecules expressed by these cells by Real-Time RT-PCR and immunohistochemical analyses. The data obtained with this work showed that this biomaterial is able to reduce the expression of some catabolic molecules by human chondrocytes and provide a good environment to support the differentiation of mesenchymal stem cells in chondrogenic sense. These observations confirm Hyaff® -11 as a suitable scaffold both for chondrocytes and mesenchymal stem cells for the treatment of articular cartilage defects. Show more

Keywords: Human chondrocytes, mesenchymal stem cells, hyaluronan-based scaffold, chondrogenic differentiation

Citation: Biorheology, vol. 43, no. 3-4, pp. 471-480, 2006

Authors: Wendt, D. | Stroebel, S. | Jakob, M. | John, G.T. | Martin, I.

Article Type: Research Article

Abstract: In this work, we assessed whether culture of uniformly seeded chondrocytes under direct perfusion, which supplies the cells with normoxic oxygen levels, can maintain a uniform distribution of viable cells throughout porous scaffolds several milimeters in thickness, and support the development of uniform tissue grafts. An integrated bioreactor system was first developed to streamline the steps of perfusion cell seeding of porous scaffolds and perfusion culture of the cell-seeded scaffolds. Oxygen tensions in perfused constructs were monitored by in-line oxygen sensors incorporated at the construct inlet and outlet. Adult human articular chondrocytes were perfusion-seeded into 4.5 mm thick foam scaffolds …at a rate of 1 mm/s. Cell-seeded foams were then either cultured statically in dishes or further cultured under perfusion at a rate of 100 μm/s for 2 weeks. Following perfusion seeding, viable cells were uniformly distributed throughout the foams. Constructs subsequently cultured statically were highly heterogeneous, with cells and matrix concentrated at the construct periphery. In contrast, constructs cultured under perfusion were highly homogeneous, with uniform distributions of cells and matrix. Oxygen tensions of the perfused medium were maintained near normoxic levels (inlet ≅ 20%, outlet > 15%) at all times of culture. We have demonstrated that perfusion culture of cells seeded uniformly within porous scaffolds, at a flow rate maintaining a homogeneous oxygen supply, supports the development of uniform engineering tissue grafts of clinically relevant thicknesses. Show more

Keywords: Bioreactor, fluid flow, chondrocyte, mass transport, functional tissue engineering

Citation: Biorheology, vol. 43, no. 3-4, pp. 481-488, 2006

Authors: Yang, Jing-Wei | Isla, Natalia de | Huselstein, Céline | Sarda-Kolopp, Marie-Nathalie | Li, Na | Li, Yin-Ping | Jing-Ping, Ou-Yang | Stoltz, Jean-François | Eljaafari, Assia

Article Type: Research Article

Abstract: Mesenchymal stem cells (MSCs) have the potential to differentiate into distinct mesenchymal tissue cells. They are easy to expand while maintaining their undifferentiated state, which suggests that these cells could be an attractive cell source for tissue engineering of cartilage. In vitro high density micromass culture has been widely used for chondrogenesis induction. Our objective was to investigate human MSCs cell cycle, viability and differentiation in these conditions. Therefore, to induce human MSCs chondrogenesis, micromasses were cultured in the presence of transforming growth factor-β1 in serum free medium for 21 days. Cell cycle, cell viability and cell phenotype were analyzed …by flow cytometry. From day 0 to 7, the G0/G1 phase increased, whereas the S phase decreased gradually, but cell cycle phases (S, G0/G1 and G2/M) did not significantly change after day 7. Less than 10% of cells were apoptotic, but no necrosis was observed, even at day 21. We observed a decrease in CD90 and CD105 expression, from day 0 to 21. In conclusion, our results demonstrate a good viability of human MSCs in micromass culture during the whole period of culture. Moreover, micromass culture allowed human MSCs to be synchronized at the G0/G1 phase, while their phenotype suggested some degree of differentiation. Show more

Keywords: Mesenchymal stem cell, TGF-β1, viability, cell cycle, differentiation

Citation: Biorheology, vol. 43, no. 3-4, pp. 489-496, 2006

Authors: Ng, Kenneth W. | Mauck, Robert L. | Statman, Lauren Y. | Lin, Evan Y. | Ateshian, Gerard A. | Hung, Clark T.

Article Type: Research Article

Abstract: The application of dynamic physiologic loading to a bilayered chondrocyte-seeded agarose construct with a 2% (wt/vol) top layer and 3% (wt/vol) bottom layer was hypothesized to (1) improve overall construct properties and (2) result in a tissue that mimics the mechanical inhomogeneity of native cartilage. Dynamic loading over the 28 day culture period was found to significantly increase bulk mechanical and biochemical properties versus free-swelling culture. The initial depth-distribution of the compressive Young's modulus (EY ) reflected the intrinsic properties of the gel in each layer and a similar trend to the native tissue, with a softer 2% gel layer …and a much stiffer 3% gel layer. After 28 days in culture, free-swelling conditions maintained this general trend while loaded constructs possessed a reverse profile, with significant increases in EY observed only in the 2% gel. Histological analysis revealed preferential matrix formation in the 2% agarose layer, with matrix localized more pericellularly in the 3% agarose layer. Finite element modeling revealed that, prior to significant matrix elaboration, the 2% layer experiences increased mechanical stimuli (fluid flow and compressive strain) during loading that may enhance chondrocyte stimulation and nutrient transport in that layer, consistent with experimental observations. From these results, we conclude that due to the limitations in 3% agarose, the use of this type of bilayered construct to construct depth-dependent inhomogeneity similar to the native tissue is not likely to be successful under long-term culture conditions. Our study underscores the importance of other physical properties of the scaffold that may have a greater influence on interconnected tissue formation than intrinsic scaffold stiffness. Show more

Keywords: Cartilage, tissue engineering, inhomogeneity, digital image correlation, agarose, scaffold design

Citation: Biorheology, vol. 43, no. 3-4, pp. 497-507, 2006

Authors: Jakab, Karoly | Damon, Brook | Neagu, Adrian | Kachurin, Anatolij | Forgacs, Gabor

Article Type: Research Article

Abstract: Bioprinting is an evolving tissue engineering technology. It utilizes computer controlled three-dimensional printers for rapid and high-precision construction of three-dimensional biological structures. We employed discrete and continuous bioprinting to build three-dimensional tissue constructs. In the former case bioink particles – spherical cell aggregates composed of many thousands of cells – are delivered one by one into biocompatible scaffolds, the biopaper. Structure formation takes place by the subsequent fusion of the bioink particles due to their liquid-like and self-assembly properties. In the latter case a mixture of cells and scaffold material is extruded from the biocartridge akin to toothpaste to arrive …at the desired construct. Specifically, we built rectangular tissue blocks of several hundred microns in thickness as well as tubular structures of several millimeters in height. The physical basis of structure formation was studied by computer simulations. Show more

Keywords: Tissue engineering, bioprinter, spherical cell aggregate, tissue liquidity, self-assembly

Citation: Biorheology, vol. 43, no. 3-4, pp. 509-513, 2006

Authors: Martin, J.A. | Buckwalter, J.A.

Article Type: Research Article

Abstract: Post-traumatic osteoarthritis is the form of osteoarthritis (OA) that develops following joint injury. Although its end-stage is indistinguishable from idiopathic OA, many patients with post-traumatic OA are younger than those with idiopathic OA, and they have a well-defined precipitating insult. Clinical and experimental studies suggest that excessive acute impact energy or chronic mechanical overload cause the degeneration of the articular surface responsible for post-traumatic OA. Yet, the mechanisms by which excessive mechanical force causes OA remain unknown. For these reasons it has not been possible to develop effective methods of preventing or decreasing the risk of post-traumatic OA. We hypothesized …that mechanical loading that exceeds the tolerance of the articular surface causes chondrocyte damage due to oxidative stress. Our in vitro tests of human articular cartilage samples showed that shear stress causes chondrocyte death and that anti-oxidants decrease the shear stress induced cell death. These observations suggest that specific patterns of loading are particularly damaging to articular surfaces and that improved treatments of joint injuries may include mechanical methods of minimizing shear stresses and biologic methods of minimizing oxidative damage. Show more

Keywords: Osteoarthritis, trauma, post-traumatic osteoarthritis, oxidative damage, chondrocytes, anti-oxidants

Citation: Biorheology, vol. 43, no. 3-4, pp. 517-521, 2006

Authors: Nieminen, Heikki J. | Töyräs, Juha | Laasanen, Mikko S. | Jurvelin, Jukka S.

Article Type: Research Article

Abstract: Mechano-acoustic and elastographic techniques may provide quantitative means for the in vivo diagnostics of articular cartilage. These techniques assume that sound speed does not change during tissue loading. As articular cartilage shows volumetric changes during compression, acoustic properties of cartilage may change affecting the validity of mechano-acoustic measurements. In this study, we examined the ultrasound propagation through human, bovine and porcine articular cartilage during stress-relaxation in unconfined compression. The time of flight (TOF) technique with known cartilage thickness (true sound speed) as well as in situ calibration method [Suh, Youn, Fu, J. Biomech. 34 (2001), 1347–1353] were used for the …determination of sound speed. Ultrasound speed and attenuation decreased in articular cartilage during ramp compression, but returned towards the level of original values during relaxation. Variations in ultrasound speed induced an error in strain and compressive moduli provided that constant ultrasound speed and time-of-flight data was used to determine the tissue thickness. Highest errors in strain (−11.8±12.0%) and dynamic modulus (15.4±17.9%) were recorded in bovine cartilage. TOF and in situ calibration methods yielded different results for changes in sound speed during compression. We speculate that the variations in acoustic properties in loaded cartilage are related to rearrangement of the interstitial matrix, especially to that of collagen fibers. In human cartilage the changes, are, however relatively small and, according to the numerical simulations, mechano-acoustic techniques that assume constant acoustic properties for the cartilage will not be significantly impaired by this phenomenon. Show more

Keywords: Articular cartilage, osteoarthrosis, mechanical indentation, ultrasound speed, ultrasound attenuation

Citation: Biorheology, vol. 43, no. 3-4, pp. 523-535, 2006

Authors: Adams, Michael A.

Article Type: Research Article

Abstract: There is a growing literature concerning chondrocyte responses to mechanical loading, but relatively little is known about the mechanical environment these cells experience in a living joint. Calculations indicate that high forces are applied to limb joints whenever the joints are flexed, because flexion can cause body weight to act on long lever arms compared to the joint centre, whereas the muscles which extend the joint act on much shorter lever arms. As a result, joint reaction forces (which compress the cartilage) can rise to 3–6 times body weight during activities such as stair climbing. Articular cartilage tends to spread …this load evenly over the joint surface, but is too thin to do this well, and compressive stresses can rise to 10–20 MPa. Within cartilage, matrix stresses vary locally, possibly as a result of variation in composition or undulations in the subchondral bone, and further modifications of stress occur within each chondron. Articular cartilage is a fibrous solid and cells within it are deformed by mechanical loading rather than subjected to a hydrostatic pressure. The mechanical environment of chondrocytes can best be reproduced in vitro by direct compression of the articular surface of cartilage which is supported naturally by adjacent cartilage and subchondral bone. Show more

Keywords: Cartilage, mechanics, mechanobiology, matrix, stress

Citation: Biorheology, vol. 43, no. 3-4, pp. 537-545, 2006

Authors: Goebel, Jean Christophe | Watrin-Pinzano, Astrid | Bettembourg-Brault, Isabelle | Odille, Freddy | Felblinger, Jacques | Chary-Valckenaere, Isabelle | Netter, Patrick | Blum, Alain | Gillet, Pierre | Loeuille, Damien

Article Type: Research Article

Abstract: Objectives: As the early form of OA is characterized by elevated water content in the cartilage tissue, the purpose of this study was to verify in vivo if age-related changes in patellar cartilage in healthy volunteers can be detected using quantitative MRI with T2 mapping and volume measurement MRI methods. Design: Thirty healthy volunteers of various classes of age (18 to 65 years old) were enrolled in this study. MR images of the patellar cartilage were acquired at 1.5T. Patellar cartilage volume and T2 maps were determined. Results: Despite non-significance, there was a trend in reducing cartilage volume with ageing …(r: −0.25). In contrast global T2 slightly increased with ageing (r: 0.46). BMI (r: 0.51) and bone volume (r: 0.69) are well correlated to cartilage volume. Conclusion. Age-related physiologic changes in the water content of patellar cartilage can be detected using MRI. The proposed T2-mapping method, coupled with other non-invasive MR cartilage imaging techniques, could aid in the early diagnosis of OA. Show more

Keywords: Cartilage, MRI, T2 map, volume, patella, age-related changes

Citation: Biorheology, vol. 43, no. 3-4, pp. 547-551, 2006

Authors: Marsano, A. | Wendt, D. | Quinn, T.M. | Sims, T.J. | Farhadi, J. | Jakob, M. | Heberer, M. | Martin, I.

Article Type: Research Article

Abstract: In this study, we aimed at validating a rotary cell culture system (RCCS) bioreactor with medium recirculation and external oxygenation, for cartilage tissue engineering. Primary bovine and human culture-expanded chondrocytes were seeded into non-woven meshes of esterified hyaluronan (HYAFF® -11), and the resulting constructs were cultured statically or in the RCCS, in the presence of insulin and TGFβ3, for up to 4 weeks. Culture in the RCCS did not induce significant differences in the contents of glycosaminoglycans (GAG) and collagen deposited, but markedly affected their distribution. In contrast to statically grown tissues, engineered cartilage cultured in the RCCS had a …bi-zonal structure, consisting of an outgrowing fibrous capsule deficient in GAG and rich in collagen, and an inner region more positively stained for GAG. Structurally, trends were similar using primary bovine or expanded human chondrocytes, although the human cells deposited inferior amounts of matrix. The use of the presented RCCS, in conjunction with the described medium composition, has the potential to generate bi-zonal tissues with features qualitatively resembling the native meniscus. Show more

Keywords: Tissue engineering, bioreactor, hydrodynamic flow, articular chondrocytes, rotating wall vessel, meniscus

Citation: Biorheology, vol. 43, no. 3-4, pp. 553-560, 2006

Authors: Moulin, David | Poleni, Paul-Emile | Kirchmeyer, Mélanie | Sebillaud, Sylvie | Koufany, Meriem | Netter, Patrick | Terlain, Bernard | Bianchi, Arnaud | Jouzeau, Jean-Yves

Article Type: Research Article

Abstract: In response to inflammatory cytokines, chondrocytes and synovial fibroblasts produce high amounts of prostaglandins (PG) which self-perpetuate locally the inflammatory reaction. Prostaglandins act primarily through membrane receptors coupled to G proteins but also bind to nuclear Peroxisome Proliferator-Activated Receptors (PPARs). Amongst fatty acids, the cyclopentenone metabolite of PGD2 , 15-deoxy-$\Delta\tsup{12,14}$ PGJ2 (15d-PGJ2 ), was shown to be a potent ligand of the PPARγ isotype prone to inhibit the production of inflammatory mediators. As the stimulated synthesis of PGE2 originates from the preferential coupling of inducible enzymes, cyclooxygenase-2 (COX-2) and membrane PGE synthase-1 (mPGES-1), we investigated the potency …of 15d-PGJ2 to regulate prostaglandins synthesis in rat chondrocytes stimulated with interleukin-1β (IL-1β). We demonstrated that 15d-PGJ2 , but not the high-affinity PPARγ ligand rosiglitazone, decreased almost completely PGE2 synthesis and mPGES-1 expression. The inhibitory potency of 15d-PGJ2 was unaffected by changes in PPARγ expression and resulted from inhibition of NF-κB nuclear binding and IκBα sparing, secondary to reduced phosphorylation of IKKβ. Consistently with 15d-PGJ2 being a putative endogenous regulator of the inflammatory reaction if synthesized in sufficient amounts, the present data confirm the variable PPARγ-dependency of its effects in joint cells while underlining possible species and cell types specificities. Show more

Keywords: PPARγ, eicosanoids, 15-deoxy-$\Delta^{12,14}$-prostaglandin J$_{2}$, thiazolidinediones, chondrocyte, synoviocyte

Citation: Biorheology, vol. 43, no. 3-4, pp. 561-575, 2006

Authors: Domagala, F. | Martin, G. | Bogdanowicz, P. | Ficheux, H. | Pujol, J.-P.

Article Type: Research Article

Abstract: In the present report we have shown that bovine articular chondrocytes cultured in low oxygen tension, i.e. in conditions mimicking their hypoxic in vivo environment, respond to IL-1β (10 ng/ml) by an increased DNA binding activity of NF-κB and AP-l transcription factors. Incubation of the cells with 10−5 M Rhein, the active metabolite of Diacerhein, for 24 h was found to reduce this activity particularly in the case of AP-1. Mitogen activated kinases (ERK-1 and ERK-2) were activated by exposure of the chondrocytes to a 1 h treatment with IL-1β. This effect was greater in hypoxia (3% O2 ) …than in normoxia (21% O2 ). Rhein was capable of reducing the IL-1β-stimulated ERK1/ERK2 pathway whatever the tension of oxygen present in the environment. The mRNA steady-state levels of collagen type II (COL2A1) and aggrecan core protein were found to be significantly increased by a 24-h treatment with 10−5 M Rhein. This stimulating effect was also observed in the presence of IL-1β, suggesting that the drug could prevent or reduce the IL-1β-induced inhibition of extra cellular matrix synthesis. IL-1-induced collagenase (MMP1) expression was significantly decreased by Rhein under the same conditions. In conclusion, Rhein can effectively inhibit the IL-1-activated MAPK pathway and the binding of NF-κB and AP-1 transcription factors, two key factors involved in the expression of several pro-inflammatory genes by chondrocytes. In addition, the drug can reduce the procatabolic effect of the cytokine, by reducing the MMP1 synthesis, and enhance the synthesis of matrix components, such as type II collagen and aggrecan. These results may explain the anti-osteoarthritic properties of Rhein and its disease-modifying effects on OA cartilage, in spite of the absence of inhibition at prostaglandin level. Show more

Citation: Biorheology, vol. 43, no. 3-4, pp. 577-587, 2006

Authors: Brandt, Kenneth D.

Article Type: Research Article

Citation: Biorheology, vol. 43, no. 3-4, pp. 589-594, 2006

Authors: de Isla, N.G. | Yang, J.W. | Huselstein, C. | Muller, S. | Stoltz, J.F.

Article Type: Research Article

Abstract: Several factors are known to be involved in the destruction of the articular cartilage. Interleukin-1 (IL-1) plays an important role in the pathogenesis of osteoarthritis (OA) either directly or through the stimulation of catabolic factors. The action of IL-1 on articular cartilage is multifaceted and it most likely plays an important role in the mechanism of cartilage destruction. IL-1 suppresses the synthesis of the cartilage matrix components and promotes the degradation of cartilage matrix macromolecules. Diacerein is an anthraquinone molecule that has been shown to reduce the severity of OA, both in man and in animal models. The present study …was designed to evaluate in vitro effects of diacerein on IL-1β expression in LPS or IL-1α stimulated chondrocytes. Intracellular IL-1β production was analysed in articular chondrocytes cultured in monolayer or in alginate 3D-biosystems in the presence of lipopolysaccharide (LPS) or IL-1α, with or without diacerein. The results show that LPS and IL-1α increase intracellular IL-1β and Diacerein inhibited LPS-induced and IL-1α induced IL-1β production by articular chondrocytes. Moreover, the effect of mechanical stimulation was analysed. An inhibitory effect of DAR at therapeutic concentrations on IL-1β production in articular chondrocytes is suggested. Show more

Keywords: Articular chondrocyte, interleukin-1β, diacerein, mechanical stress, osteoarthritis

Citation: Biorheology, vol. 43, no. 3-4, pp. 595-601, 2006

Authors: Buckwalter, Joseph A. | Martin, James A. | Brown, Thomas D.

Article Type: Research Article

Abstract: Osteoarthritis, the clinical syndrome of joint pain and dysfunction due to joint degeneration, is among the most frequent and symptomatic medical problems for middle aged and older people, and it is the most common cause of long term disability in most populations of people over 65. Currently there are no effective methods of preventing or curing osteoarthritis. Post-traumatic OA, the joint degeneration, pain and dysfunction that develop following joint injury, is the form of OA that is most directly related to elevated articular surface contact stress. However, mechanical stress that exceeds the tolerance of the articular surface can cause or …accelerate the progression of joint degeneration in all individuals and in all synovial joints. In some patients, decreasing mechanical forces on degenerated joint surfaces stimulates formation of a new biologic articular surface. The advances in understanding of the effects of mechanical forces on chondrocytes and cartilage presented and discussed at the 4th Symposium on Mechanobiology: Cartilage and Chondrocyte will help in the efforts to develop new methods of preventing and treating osteoarthritis. Show more

Citation: Biorheology, vol. 43, no. 3-4, pp. 603-609, 2006