Cartilage Disc Research Articles

Bioprinting Tissue Analogues with Decellularized Extracellular Matrix Bioink for Regeneration and Tissue Models of Cartilage and Intervertebral Discs

AbstractThe recent convergence of decellularized extracellular matrix (dECM) methodologies and 3D bioprinting (3DBP) has led to multiple advancements in tissue engineering scaffold design by enabling researchers to recreate a tissue‐specific 3D environment in a custom geometry. The application of 3DBP of dECM for articular cartilage and intervertebral disc (IVD) repair, however, is still burgeoning. While cartilage and IVD tissue each possess unique architectures, they are composed of similar macromolecules and therefore pose similar challenges for the successful application of their matrix bioinks. Herein, the state‐of‐the‐art in cartilage and IVD dECM bioink preparation, material properties, and applications are highlighted. The current major obstacles regarding optimal decellularization and printing methods are discussed, which need to be overcome to enable recapitulation of the hierarchical organization, zone‐specific matrix composition, and anisotropic biomechanical behavior of the native tissues. Finally, a vision is presented for how this field may continue to evolve in the future to empower the fabrication of scaffolds that serve as effective templates for guiding cellular assembly and organization toward the formation of functional cartilage and IVD tissues.

FAKTOR-FAKTOR YANG BERHUBUNGAN DENGAN KEJADIAN MUSCULOSKELETAL DISORDERS (MSDs) PADA PEMANEN KELAPA SAWIT DI PT. JOHAN SENTOSA

The European Commission Supervisory Report calculates cases of MSDs causing 49.9% absence from work for more than three days and 60% permanent disability at work. While in Korea, MSD reduced a very high increase from 1,634 in 2001 to 5,502 in 2010. Musculoskeletal disorders (MSDs) are damage to muscles, nerves, tendons, ligaments, joints, cartilage or spinal discs. The purpose of this study was to study the factors associated with the occurrence of musculoskeletal disorders (MSDs) in oil palm harvesters at PT Johan Sentosa Bangkinang in 2019. This type of research is the type of this research is analytical research using cross sectional surveys. The population of this research was all 93 palm oil harvesters with 75 samples. The sampling technique uses simple random sampling. Data aids using a questionnaire. Data analysis in this study used univariate and bivariate analysis. The research results obtained from there is a significant relationship between Age and MSDs incidence p value 0,002 RP = 1,805 (95% CI: 1,224-2,661), there is a significant relationship between Work Periods and Event MSDs p value 0,001 Rp = 1,951 (CI: 95 %: 1,260-3,020), there is no relationship involving weight and height with the incidence of MSDs p value 0,314), there is a significant relationship between the Age of Oil Palm with the incidence of MSDs p value 0.001 RP = 1,944 (95% CI: 1,232-3,067 ) and there is a significant relationship between Land Contour and MSDs p value 0.003 RP = 1.738 (95% CI: 1.94-2.529). For PT. Johan Sentosa is expected to be able to provide knowledge about how to work really when harvesting so workers can avoid the MSD incident.

Cell-Laden Agarose-Collagen Composite Hydrogels for Mechanotransduction Studies.

The increasing investigation of cellular mechanotransduction mechanisms requires biomaterials combining biofunctionality and suitable mechanical properties. Agarose is a standard biomaterial for cartilage and intervertebral disc mechanobiology studies, but lacks adhesion motifs and the necessary cell-matrix interaction for mechanotransduction. Here, collagen type I was blended at two concentrations (2 and 4.5 mg/mL) with agarose 2% wt/vol. The composite hydrogels were characterized in terms of structural homogeneity, rheological properties and size stability. Nucleus pulposus (NP) cell viability, proliferation, morphology, gene expression, GAG production, adhesion and mechanotransduction ability were further tested. Blended hydrogels presented a homogenous network of the two polymers. While the addition of 4.5 mg/mL collagen significantly decreased the storage modulus and increased the loss modulus of the gels, blended gels containing 2 mg/mL collagen displayed similar mechanical properties to agarose. Hydrogel size was conserved over 21 days for all agarose-based gels. Embedded cells were viable (>80%) and presented reduced proliferation and a round morphology typical of NP cells in vivo. Gene expression of collagen types I and II and aggrecan significantly increased in blended hydrogels from day 1 to 7, further resulting in a significantly superior GAG/DNA ratio compared to agarose gels at day 7. Agarose-collagen hydrogels not only promoted cell adhesion, contrary to agarose gels, but also showed a 5.36-fold higher focal adhesion kinase phosphorylation (pFAK/β-tubulin) when not compressed, and increased pFAK/FAK values 10 min after compression. Agarose-collagen thus outperforms agarose, mimics native tissues constituted of non-fibrillar matrix and collagens, and allows exploring complex loading in a highly reproducible system.

Functional Deficits in Mice Expressing Human Interleukin 8.

We showed previously that inflammatory mediators, including IL8, in intervertebral disc tissues from patients with discogenic back pain may play a key role in back pain. To investigate the molecular mechanism of IL8 signaling in back pain, we generated a mouse model that conditionally expresses human (h) IL8. We hypothesized that hIL8 levels affect mouse activity and function. Briefly, hIL8 cDNA was inserted into the pCALL2 plasmid, linearized, and injected into mouse embryos. Resulting pCALL2-hIL8 mice were then bred with GDF5-Cre mice to express the transgene in cartilage and intervertebral disc (IVD) tissues. Functional capacities including nest-making and other natural behaviors were measured. Both male and female mice expressing hIL8 showed lower nesting scores than did littermates that did not express hIL8 (n = 14 to 16 per group). At 28 wk of age, mice expressing hIL8 (n = 35) spent more time immobile and eating during each night than littermate controls (n = 33). Furthermore, hIL8-expressing mice traveled shorter distances and at a lower average speed than littermate controls. Thus, in an initial effort to investigate the relationship between this chemokine and mouse behavior, we have documented changes in normal activities in mice conditionally expressing hIL8.

The Intra-Articular Hematoma Immediately after Ankle Fracture Causes Cartilage Damage That is Partially Attenuated by Anti-Inflammatory Agents

Category:Ankle Arthritis; Basic Sciences/BiologicsIntroduction/Purpose:Post-traumatic osteoarthritis (PTOA) is a frequent cause of disability. The most common predisposing factor for ankle PTOA is intra-articular ankle fracture. It has been hypothesized that an early inflammatory response after intra- articular injury could lead to cartilage damage. Our group recently demonstrated significantly elevated inflammatory cytokines in the synovial fluid fracture hematoma (SFFH) immediately after intra-articular ankle fracture. The negative effect that this inflammatory environment has on cartilage is unknown but may be the reason for PTOA development. The purpose of this study was to determine if cartilage occurred when exposed to intra-articular SFFH and if anti-inflammatory agents could attenuate this damage.Methods:SFFH was obtained from 52 intra-articular ankle fractures at the time of surgery. Samples were divided into three groups based on time from fracture: acute (0-2 days), intermediate (3-9 days), late (>=10 days). A biopsy punch was used to create 5mm cartilage discs from the medial and lateral shoulders of fresh human talus cartilage. The cartilage discs were weighed and cultured in standard chondrocyte media (SCM) for 3 days. Next, the discs were randomly assigned to one of 4 groups (n=10 each) and cultured as follows for 6 days: A=control, SCM only; Group B=acute SFFH+SCM; Group C=intermediate SFFH+SCM; Group D=late SFFH+SCM. After 6 days of culture, the discs were rinsed and transferred to new wells and cultured for 3 more days in SCM only, to determine inflammatory cytokine and MMP release from the cartilage discs (cartilage damage). The post SFFH spent culture media was collected. The initial and post SFFH culture spent media were analyzed for inflammatory cytokines, MMPs, CTXII (cartilage breakdown marker), and sGAG. Safranin-O staining was then performed on the discs. The initial and post SFFH media from groups A, B, C, and D were compared to determine if there was a difference in damage caused by SFFH based on time from fracture. Based on the results, Group B (early) was felt to cause the most damage. Therefore, the following treatment groups were created and cultured exactly as before: Group E=same as Group B + IL-1Ra treatment; Group F=same as Group B + doxycycline treatment. Groups B, E, and F were compared to determine if the addition of IL-1Ra or doxycycline (known MMP inhibitor) attenuated the damage.Results:Early, intermediate, and late SFFH all caused some degree of cartilage damage in the form of significantly elevated inflammatory cytokine and MMP production from the cartilage discs (Figure 1). The early SFFH (Group B) was noted to cause significantly elevated production of IL-6, IL-8, CTXII, and decreased safranin-O staining. Therefore, it was chosen to compare with the therapeutic anti-inflammatory agents. The addition of IL-1Ra or doxycycline significantly reduced the production of IL-6, IL-8, CTXII, and MMPs 3, 9, and 10 (Figure 2).Conclusion:This is the first study to show that intra-articular hematoma immediately after ankle fracture causes cartilage damage that can be partially attenuated by current clinically available therapeutic agents. Additionally, further research should be performed regarding the safety and efficacy of anti-inflammatory agents to prevent ankle PTOA.

Three-Dimensional Bioprinting in Orthopaedics.

Three-dimensional (3D)-printing technology has evolved dramatically in the last 30 years, from large machines with poor resolution to those with micron-level capabilities that sit on a desktop. This technology is being utilized in numerous medical applications, particularly in orthopaedic surgery. Over the past decade, technological advances have allowed for the application of this technology to the field of tissue engineering through the process of 3D bioprinting. Of interest to orthopaedic surgeons, active areas of research utilizing this technology involve the bioprinting of articular cartilage, bone, menisci, and intervertebral discs.

Relationship between Facet Joint Asymmetry and Adjacent Intervertebral Disc Degeneration in Lumbar Disc Herniation

Objective: To analyze the relationship between facet joint asymmetry and adjacent intervertebral disc degeneration in lumbar disc herniation. Methods: Fifty patients with L4/5 lumbar disc herniation were enrolled in the study. All patients underwent lumbar MRI examination, to compare the facet asymmetry of lumbar disc herniation and the degeneration of cartilage endplate and intervertebral disc of adjacent segments, and analyze the relationship between them. Results: There was no significant correlation between L4/5 facet asymmetry and L5/S1 intervertebral disc degeneration score and cartilage endplate degeneration score (P > 0.05). L4/5 facet asymmetry was significantly correlated with L3/4 disc degeneration score (P P > 0.05). Conclusion: There was a positive correlation between the facet joint angle of lumbar disc herniation and the degeneration of upper proximal segment intervertebral disc. Early correction of lumbar facet angle asymmetry can provide beneficial guidance for further prevention and treatment.

The development of synthetic analogues of glycosaminoglycans for the cost-effective modification of contact lens surfaces

Purpose Medical devices benefit from increasingly sophisticated mimics of bodily tissues; contact lenses are no exception and additionally rely on cost-effective process technology. Bodily hydration is often regulated by sulfated glycosaminoglycans (GAGs, e,g. chondroitin sulfate). Hydration is particularly important in dermal, ocular and orthopaedic (cartilage and intervertebral disc) applications. This poster describes synthetic approaches to harness synthetic GAG analogues in the cost-effective modification of contact lens surfaces.

Articulation of an alumina-zirconia composite ceramic against living cartilage – An in vitro wear test

BackgroundThere is interest in minimally invasive solutions that reduce osteoarthritic symptoms and restore joint mobility in the early stages of cartilage degeneration. The aim of the present study was to evaluate an alumina-zirconia composite (AZC) as a counterface for articulation against live cartilage explants in comparison to the clinically relevant cobalt-chromium (CoCrMo) alloy. MethodsA four-station bioreactor designed to articulate against living tissue in an incubator was used for testing. Twelve 32 mm AZC and twelve 32 mm CoCrMo femoral heads with equal surface roughness made up both test groups. Each head articulated against a cartilage disk harvested from stifle joints of 24-week old steers. Test samples and free-swelling control cartilage samples were cultured in Mini ITS medium. Testing was conducted 3 h daily over 10 days applying a contact load of approximately 2 MPa. PG/GAG and hydroxyproline were analyzed using biochemical assays. Additionally, chondrocyte survival and Mankin score analyses were performed on histological slides. ResultsCells stayed alive during the course of the experiment, with cell survival values close to 80% at test completion in the superficial zone. There was no significant difference between AZC and free-swelling control tissue. However, cell count values were inferior for CoCrMo (p = 0.003). Tested tissue suffered mostly structural abnormalities. The PG/GAG content in medium was not different between CoCrMo and AZC (p = 0.315); however, the hydroxyproline release into medium was nearly 30% higher for CoCrMo (p = 0.024). ConclusionBased on average values, AZC induces less cell and tissue damage than CoCrMo. However, only the hydroxyproline measurements reached statistical significance, partially due to substantial scatter within both groups.

Sirtuin 2 Promotes Collagen II Synthesis Through Deacetylating p65-NF-κB in Human Intervertebral Disc Cartilage Endplate Cells

Backgrounds: Intervertebral disc degeneration (IDD) is a major cause of low back pain, however its underlying pathogenesis is complicated and remains extensively illumination. Here we investigate the role of sirtuin 2 (SIRT2) in human IDD. Methods: We compared the RNA and protein levels of SIRT2 in non-degenerate and degenerate human tissues and assessed their relationship with clinical features. After isolated the intervertebral disc cartilage endplate (CEP) cells, the cell proliferation and apoptosis were tested. Further underlying mechanisms were explored from molecular levels. Findings: We found that the CEP cell proliferation and apoptosis was not changed during IDD. However, the CEP collagen synthesis and secretion were attenuated in IDD patients compared with healthy individuals. Moreover, the SIRT2 expression was downregulated in the CEP cells isolated from IDD patients. Overexpressing SIRT2 can significantly rescue the capacity of collagen II synthesis. Upon SIRT2 overexpression, the NF-κB was deacetylated and subsequently modulate collagen gene transcription, therefore preventing IDD development. Interpretation: SIRT2 downregulation is one of the possible causes of IDD during aging. Overexpressing SIRT2 will attenuate IDD progression via deactivating NFκB. Funding Statement: This work is supported by the National Natural Science Foundation of China (NSFC #81601957) Declaration of Interests: The authors state: None declared. Ethics Approval Statement: The work was approved by the Ethics Committee of the China–Japan Union Hospital (Changchun, China). Written informed consent was obtained from patients or relatives before tissue collection. All procedures are complied with the principles outlined in the Declaration of Helsinki.

Exogenous Delivery of Link N mRNA into Chondrocytes and MSCs-The Potential Role in Increasing Anabolic Response.

Musculoskeletal disorders, such as osteoarthritis and intervertebral disc degeneration are causes of morbidity, which concomitantly burdens the health and social care systems worldwide, with massive costs. Link N peptide has recently been described as a novel anabolic stimulator for intervertebral disc repair. In this study, we analyzed the influence on anabolic response, by delivering synthetic Link N encoding mRNA into primary human chondrocytes and mesenchymal stromal cells (SCP1 cells). Furthermore, both cell types were seeded on knitted titanium scaffolds, and the influence of Link N peptide mRNA for possible tissue engineering applications was investigated. Synthetic modified Link N mRNA was efficiently delivered into both cell types and cell transfection resulted in an enhanced expression of aggrecan, Sox 9, and type II collagen with a decreased expression of type X collagen. Interestingly, despite increased expression of BMP2 and BMP7, BMP signaling was repressed and TGFβ signaling was boosted by Link N transfection in mesenchymal stromal cells, suggesting possible regulatory mechanisms. Thus, the exogenous delivery of Link N peptide mRNA into cells augmented an anabolic response and thereby increased extracellular matrix synthesis. Considering these findings, we suppose that the cultivation of cells on knitted titanium scaffolds and the exogenous delivery of Link N peptide mRNA into cells could mechanically support the stability of tissue-engineered constructs and improve the synthesis of extracellular matrix by seeded cells. This method can provide a potent strategy for articular cartilage and intervertebral disc regeneration.

A radical switch in clonality reveals a stem cell niche in the epiphyseal growth plate.

Longitudinal bone growth in children is sustained by growth plates, narrow discs of cartilage that provide a continuous supply of chondrocytes for endochondral ossification1. However, it remains unknown how this supply is maintained throughout childhood growth. Chondroprogenitors in the resting zone are thought to be gradually consumed as they supply cells for longitudinal growth1,2, but this model has never been proved. Here, using clonal genetic tracing with multicolour reporters and functional perturbations, we demonstrate that longitudinal growth during the fetal and neonatal periods involves depletion of chondroprogenitors, whereas later in life, coinciding with the formation of the secondary ossification centre, chondroprogenitors acquire the capacity for self-renewal, resulting in the formation of large, stable monoclonal columns of chondrocytes. Simultaneously, chondroprogenitors begin to express stem cell markers and undergo symmetric cell division. Regulation of the pool of self-renewing progenitors involves the hedgehog and mammalian target of rapamycin complex 1 (mTORC1) signalling pathways. Our findings indicate that a stem cell niche develops postnatally in the epiphyseal growth plate, which provides a continuous supply of chondrocytes over a prolonged period.

A novel mechanobiological model can predict how physiologically relevant dynamic loading causes proteoglycan loss in mechanically injured articular cartilage

Cartilage provides low-friction properties and plays an essential role in diarthrodial joints. A hydrated ground substance composed mainly of proteoglycans (PGs) and a fibrillar collagen network are the main constituents of cartilage. Unfortunately, traumatic joint loading can destroy this complex structure and produce lesions in tissue, leading later to changes in tissue composition and, ultimately, to post-traumatic osteoarthritis (PTOA). Consequently, the fixed charge density (FCD) of PGs may decrease near the lesion. However, the underlying mechanisms leading to these tissue changes are unknown. Here, knee cartilage disks from bovine calves were injuriously compressed, followed by a physiologically relevant dynamic compression for twelve days. FCD content at different follow-up time points was assessed using digital densitometry. A novel cartilage degeneration model was developed by implementing deviatoric and maximum shear strain, as well as fluid velocity controlled algorithms to simulate the FCD loss as a function of time. Predicted loss of FCD was quite uniform around the cartilage lesions when the degeneration algorithm was driven by the fluid velocity, while the deviatoric and shear strain driven mechanisms exhibited slightly discontinuous FCD loss around cracks. Our degeneration algorithm predictions fitted well with the FCD content measured from the experiments. The developed model could subsequently be applied for prediction of FCD depletion around different cartilage lesions and for suggesting optimal rehabilitation protocols.

Water dynamics in silanized hydroxypropyl methylcellulose based hydrogels designed for tissue engineering

Silanized hydroxypropyl methylcellulose based hydrogels were developed for cartilage and intervertebral disc tissue engineering. Herein, study of dynamics of confined water showed two different populations, identified as hydration and bulk-like water. The diffusion coefficient showed that bulk-like water diffuses over distances ∼10 μm without being affected by the hydrogel matrix. Addition of silica nanofibers leads to improved mechanical properties and enhanced diffusion coefficient. Good diffusion within hydrogels is essential for the application.

Foxp2 regulates anatomical features that may be relevant for vocal behaviors and bipedal locomotion

Fundamental human traits, such as language and bipedalism, are associated with a range of anatomical adaptations in craniofacial shaping and skeletal remodeling. However, it is unclear how such morphological features arose during hominin evolution. FOXP2 is a brain-expressed transcription factor implicated in a rare disorder involving speech apraxia and language impairments. Analysis of its evolutionary history suggests that this gene may have contributed to the emergence of proficient spoken language. In the present study, through analyses of skeleton-specific knockout mice, we identified roles of Foxp2 in skull shaping and bone remodeling. Selective ablation of Foxp2 in cartilage disrupted pup vocalizations in a similar way to that of global Foxp2 mutants, which may be due to pleiotropic effects on craniofacial morphogenesis. Our findings also indicate that Foxp2 helps to regulate strength and length of hind limbs and maintenance of joint cartilage and intervertebral discs, which are all anatomical features that are susceptible to adaptations for bipedal locomotion. In light of the known roles of Foxp2 in brain circuits that are important for motor skills and spoken language, we suggest that this gene may have been well placed to contribute to coevolution of neural and anatomical adaptations related to speech and bipedal locomotion.

Effects of spaceflight on cartilage: implications on spinal physiology.

Spaceflight alters normal physiology of cells and tissues observed on Earth. The effects of spaceflight on the musculoskeletal system have been thoroughly studied, however, the effects on cartilage have not. This area is gaining more relevance as long duration missions, such as Mars, are planned. The impact on intervertebral discs and articular cartilage are of particular interest to astronauts and their physicians. This review surveys the literature and reports on the current body of knowledge regarding the effects of spaceflight on cartilage, and specifically changes to the spine and intervertebral disc integrity and physiology. A systematic literature review was conducted using PubMed, MEDLINE, and Google Scholar. Eighty-six unique papers were identified, 15 of which were included. The effect of spaceflight on cartilage is comprehensively presented due to limited research on the effect of microgravity on the spine/intervertebral discs. Cellular, animal, and human studies are discussed, focusing on human physiologic changes, cartilage histology, mineralization, biomechanics, chondrogenesis, and tissue engineering. Several common themes were found, such as decreased structural integrity of intervertebral disks and impaired osteogenesis/ossification. However, studies also presented conflicting results, rendering strong conclusions difficult. The paucity of human cartilage studies in spaceflight leaves extrapolation from other model systems the only current option for drawing conclusions despite known and unknown limitations in applicability to human physiology, especially spinal pathophysiology which is special interest. The aerospace and biomedical research communities would benefit from further human spaceflight articular cartilage and intervertebral disc studies. Further research may yield beneficial application for spaceflight, and crossover in understanding and treating terrestrial diseases like osteoarthritis and vertebral disc degeneration.

Cartilage Endplate Thickness Variation Measured by Ultrashort Echo-Time MRI Is Associated With Adjacent Disc Degeneration.

A magnetic resonance imaging study of human cadaver spines. To investigate associations between cartilage endplate (CEP) thickness and disc degeneration. Damage to the CEP is associated with spinal injury and back pain. However, CEP morphology and its association with disc degeneration have not been well characterized. Ten lumbar motion segments with varying degrees of disc degeneration were harvested from six cadaveric spines and scanned with magnetic resonance imaging in the sagittal plane using a T2-weighted two-dimensional (2D) sequence, a three-dimensional (3D) ultrashort echo-time (UTE) imaging sequence, and a 3D T1ρ mapping sequence. CEP thicknesses were calculated from 3D UTE image data using a custom, automated algorithm, and these values were validated against histology measurements. Pfirrmann grades and T1ρ values in the disc were assessed and correlated with CEP thickness. The mean CEP thickness calculated from UTE images was 0.74 ± 0.04 mm. Statistical comparisons between histology and UTE-derived measurements of CEP thickness showed significant agreement, with the mean difference not significantly different from zero (P = 0.32). Within-disc variation of T1ρ (standard deviation) was significantly lower for Pfirrmann grade 4 than Pfirrmann grade 3 (P < 0.05). Within-disc variation of T1ρ and adjacent CEP thickness heterogeneity (coefficient of variation) had a significant negative correlation (r = -0.65, P = 0.04). The standard deviation of T1ρand the mean CEP thickness showed a moderate positive correlation (r = 0.40, P = 0.26). This study demonstrates that quantitative measurements of CEP thickness measured from UTE magnetic resonance imaging are associated with disc degeneration. Our results suggest that variability in CEP thickness and T1ρ, rather than their mean values, may serve as valuable diagnostic markers for disc degeneration. N/A.

Phospholipid Vesicles in Media for Tribological Studies against Live Cartilage.

Pre-clinical testing of hemiarthroplasty devices requires that the tribological conditions present in vivo with live cartilage be closely duplicated. A current limitation in the tribological testing of live cartilage involves the use of cell-culture media as lubricant. to develop and test a new hyaluronan-phospholipid based medium (HA-phospholipid medium) that combines the rheological and frictional properties of synovial fluid with the nourishing properties of culture media to keep cells alive. The HA-phospholipid medium consisted of culture medium with added phospholipid dipalmitoylphosphatidylcholine (0.3 mg/mL), and hyaluronic acid (2.42 mg/mL). A standard cell culture medium was used as the control. The rheology of each medium was determined using a flat plate configuration. Bovine calf cartilage was used to assess cell viability and friction in each medium. For friction measurements, a cobalt-chrome alloy ball was articulated against cartilage disks immersed in medium. Lipid vesicles 0.1 to 50 μm in diameter were identified in the HA-phospholipid medium. Cartilage cell viability was significantly higher in the HA-phospholipid medium (62% ± 8%, 95% CI) than in control medium (49.5% ± 5%) (p = 0.009). The HA-phospholipid medium exhibited strong shear-thinning behavior, similar to synovial fluid, with viscosities ~100-fold higher at 10 s-1 and 5-fold higher at 20,000 s-1 than the approximately Newtonian control medium. The HA-phospholipid medium also yielded 20% lower friction values than the control medium after one hour of testing. The rheological and friction results indicate that the HA-phospholipid medium is superior to the control cell culture medium in emulating the shear thinning and lubricative properties of natural synovial fluid, making it more clinically relevant for in vitro wear and friction testing with live cartilage.

BNIP3/Bcl-2-mediated apoptosis induced by cyclic tensile stretch in human cartilage endplate-derived stem cells.

The present study aimed to investigate the molecular mechanisms of cyclic stretch-induced apoptosis in human intervertebral disc cartilage endplate-derived stem cells (CESCs). CESCs were stretched by the Flexercell-4000™ Tension Plus system, the effect on cell viability was measured by a Cell Counting Kit-8 assay, while cell apoptosis was detected by flow cytometry. Western blot analysis was used to evaluate the expression of B-cell lymphoma 2 (Bcl-2)/adenovirus E1B 19 kDa interacting protein 3 (BNIP3), Bcl-2, Bcl-2 homologous antagonist killer (Bak), Bcl-2-associated X protein (Bax), Bcl extra large (Bcl-xl) and the activity of caspase-3, while Z-VAD-FMK was used to inhibit caspase-3. Compared with the control group, the cell viability decreased in a time-dependent manner after stretching. Furthermore, cell apoptosis and the activity of caspase-3 were increased in a time-dependent manner. The ratio of pro-death factor BNIP3 to anti-apoptotic protein Bcl-2 was significantly increased. When cells were stretched for 36 h, the apoptosis-associated proteins Bak and Bax were increased, while Bcl-xl was decreased. The viability and apoptotic ratio of stretched cells were significantly restored after caspase-3 was repressed. In conclusion, cyclic tensile stretch induced apoptosis of CESCs, which was probably due to upregulation of the expression of BNIP3.

External Asymmetry and Pectoral Fin Loss in the Bamboo Sole (Heteromycteris japonica): Small-Sized Sole with Potential as a Pleuronectiformes Experimental Model.

Pleuronectiform fish develop marked external asymmetry in eye location and skin color at metamorphosis. The bamboo sole, Heteromycteris japonica, also exhibits loss of the pectoral fins at metamorphosis. Because of its small body size, short generation time, and long spawning season, we focused on the bamboo sole as an experimental model to investigate metamorphic asymmetry and pectoral fin loss during development. In the present study, we utilized a small-scale culture system to evaluate bamboo sole larvae and larval development, and a microinjection system for fertilized eggs. The culture system described here uses an 18 L culture tank for rotifers (the first diet for larvae) and 5 L plastic beakers for larval culture. Under this system, most larvae completed metamorphosis, including one-eye migration and pigmentation of the ocular side, by 23 days postfertilization (dpf) at 25°C. Larvae at density of 120-150 per liter were grown from hatching to 23 dpf with a survival ratio of 60-75% per beaker. Pectoral fins, including coracoid and disk cartilage, formed but were completely lost in late metamorphosis without formation of proximal radials and fin rays. The microinjection system designed here is adequate for the bamboo sole and allows injection of 100 one-cell-stage embryos per day. We expect that the culture and microinjection systems described here will facilitate the use of the bamboo sole as an experimental model organism in developmental biology.