Cartilage Layer Research Articles

Anisotropy of T2 and T1ρ relaxation time in articular cartilage at 3 T.

The anisotropy of R2 and R1ρ relaxation rates in articular cartilage contains information about the collagenous structure of the tissue. Here we determine and study the anisotropic and isotropic components of T2 and T1ρ relaxation parameters in articular cartilage with a clinical 3T MRI device. Furthermore, a visual representation of the topographical variation in anisotropy is given via anisotropy mapping. Eight bovine stifle joints were imaged at 22 orientations with respect to the main magnetic field using T2, continuous-wave (CW) T1ρ, and adiabatic T1ρ mapping sequences. Relaxation rates were separated into isotropic and anisotropic relaxation components using a previously established relaxation anisotropy model. Pixel-wise anisotropy values were determined from the relaxation-time maps using Michelson contrast. The relaxation rates obtained from the samples displayed notable variation depending on the sample orientation, magnetization preparation, and cartilage layer. R2 demonstrated significant anisotropy, whereas CW-R1ρ (300 Hz) and CW-R1ρ (500 Hz) displayed a low degree of anisotropy. Adiabatic R1ρ was largely isotropic. In the deep cartilage regions, relaxation rates were generally faster and more anisotropic than in the cartilage closer to the tissue surface. The isotropic relaxation rate components were found to have similar values regardless of measurement sequence. The fitted relaxation model for T2 and T1ρ demonstrated varying amounts anisotropy, depending on magnetization preparation, and studied the articular cartilage layer. Anisotropy mapping of full joints showed varying amounts of anisotropy depending on the quantitative MRI parameter and topographical location, and in the case of T2, showed systematic changes in anisotropy across cartilage depth.

Mechanism of Tougu Xiaotong Capsules in delaying degeneration of osteoarthritis by regulating cholesterol metabolism in chondrocytes through lncRNA MALAT1

From the perspective of lncRNA MALAT1 regulating cholesterol metabolism in chondrocytes, this paper explores the effect and mechanism of Tougu Xiaotong Capsules(TGXTC) in delaying the degeneration of osteoarthritis. After one week of adaptive feeding, 48(8-week-old) C57BL/6 mice were randomly divided into a blank group(12 mice) and a model group(36 mice) by random number table method. The mice in the model group were anesthetized by inhalation of 5% isoflurane, and the OA model was induced by Hulth method. The experiment randomly divided the mice into a model group(12 mice), a drug-positive group(taururso-deoxycholic acid)(12 mice), and a TGXTC group(12 mice). The drug-positive group was given 500 mg·kg~(-1) taurodeoxycholic acid by intragastric administration. TGXTC group was given TGXTC 368 mg·kg~(-1) by gavage. The blank group and model group were given the same amount of normal saline for four weeks. After the intervention, the mice in each group were killed under anesthesia, and the knee cartilage tissue was separated and collected. The morphologic changes of knee cartilage were observed. The level of lncRNA MALAT1 in the cartilage tissue was detected by real-time PCR. The protein expressions of ABCA1, ApoA1, LXRβ, CHOP, and caspase-3 in mouse articular cartilage were detected by Western blot. Lentivirus-coated plasmid was used to transfect mouse chondrocytes with sh-MALAT1. The gene levels of lncRNA MALAT1 in mouse chondrocytes transfected with sh-MALAT1 were detected by real-time PCR. Western blot was used to detect the effect of TGXTC on the protein content of ABCA1, ApoA1, LXRβ, CHOP, and caspase-3 in thapsigargin(TG)-induced mouse chondrocytes after lncRNA MALAT1 knockdown. Flow cytometry was used to detect the effect of TGXTC on apoptosis of TG-induced mouse chondrocytes after lncRNA MALAT1 knockdown. The results of HE and saffranine O staining showed that compared with the model group, the structure of the cartilage layer was basically intact; the damage degree of joint structure was significantly improved, and the cartilage matrix was significantly enhanced by saffranine O staining in the TGXTC group and drug-positive group. Compared with the model group, the lncRNA MALAT1 level was significantly decreased in the TGXTC group and drug-positive group. Compared with the model group, the protein content of ABCA1, ApoA1, and LXRβ was significantly increased, while that of CHOP and caspase-3 in the TGXTC group and drug-positive group significantly decreased. Compared with the TG group, the lncRNA MALAT1 level in the TG+sh-MALAT1 group was decreased. The lncRNA MALAT1 level in the TG+sh-MA-LAT1+TGXTC group was increased compared with the TG+TGXTC group. Western blot results showed that compared with the model group, protein expressions of ABCA1, ApoA1, LXRβ, CHOP, and caspase-3 in the TGXTC group were significantly decreased, after lncRNA MALAT1 knockdown, the regulation and apoptosis of ABCA1, ApoA1, LXRβ, CHOP, and caspase-3 in TG-induced mouse chondrocytes were weakened by TGXTC. TGXTC can improve the disorder of cholesterol metabolism in OA chondrocytes and delay OA degeneration, which is closely related to the regulation of lncRNA MALAT1.

Designing Biomimetic 3D-Printed Osteochondral Scaffolds for Enhanced Load-Bearing Capacity.

Osteoarthritis is a debilitating chronic joint disorder that affects millions of people worldwide. Since palliative and surgical treatments cannot completely regenerate hyaline cartilage within the articulating joint, osteochondral (OC) tissue engineering has been explored to heal OC defects. Utilizing computational simulations and three-dimensional (3D) printing, we aimed to build rationale around fabricating OC scaffolds with enhanced biomechanics. First, computational simulations revealed that interfacial fibrils within a bilayer alter OC scaffold deformation patterns by redirecting load-induced stresses toward the top of the cartilage layer. Principal component analysis revealed that scaffolds with 800 μm long fibrils (scaffolds 8A-8H) possessed optimal biomechanical properties to withstand compression and shear forces. While compression testing indicated that OC scaffolds with 800 μm fibrils did not have greater compressive moduli than other scaffolds, interfacial shear tests indicated that scaffold 8H possessed the greatest shear strength. Lastly, failure analysis demonstrated that yielding or buckling models describe interfacial fibril failure depending on fibril slenderness S. Specifically for scaffolds with packing density n = 6 and n = 8, the yielding failure model fits experimental loads with S < 10, while the buckling model fitted scaffolds with S < 10 slenderness. The research presented provides critical insights into designing 3D printed interfacial scaffolds with refined biomechanics toward improving OC tissue engineering outcomes.

АРТРОСКОПИЧЕСКАЯ ХОЛОДНО-ПЛАЗМЕННАЯ АБЛЯЦИЯ ПРИ РЕДКИХ ЗАБОЛЕВАНИЯХ СИНОВИАЛЬНОЙ ОБОЛОЧКИ КОЛЕННОГО СУСТАВА. КЛИНИЧЕСКИЕ НАБЛЮДЕНИЯ

Background. Over a long period, an open capsular synovectomy technique was recommended for villous nodular synovitis and synovial chondromatosis of the knee joint. However, such surgical treatment is accompanied by significant trauma to the joint tissue and frequent postoperative recurrence. Currently, it is advisable to assess the possibilities of modern treatment methods (arthroscopic cold plasma ablation) for rare diseases of the knee joint. The aim of the study. To assess the possibilities of arthroscopic cold plasma ablation in a series of clinical observa-tions of the surgical treatment of patients with rare diseases of the knee joint synovium in a statistically small sample. Material and methods. Surgical treatment was performed in 14 patients with diseases of the knee joint synovium. We carried out clinical and radiological examinations, computed tomography and magnetic resonance imaging, multi-detector computed tomographic angiography, ultrasound examination, Dopplerography of the vessels of the lower extremities, biochemical examination of synovial fluid, arthroscopy with targeted biopsy and subsequent morphological examination of the synovial membrane. Results. In 7 patients with pigmented villous nodular synovitis, the following surgeries were performed: in 1 patient with the diffuse form – an open subtotal capsular sinovectomy; in 6 patients with the nodular form – an arthroscopic partial cold plasma capsular sinovectomy of the knee joint. In 7 patients with synovial chondromatosis, an arthroscopic removal of cartilage flaps and partial cold plasma ablation of synovial folds and knee joint villi were carried out. Conclusion. Arthroscopic cold plasma ablation with bipolar electrodes in various modes of the Qvantum-2 (Arthro-Care, USA) using anterior, posterolateral and suprapatellar approaches makes it possible to carry out less traumatic and radical, without damaging deep layers of cartilage and capsule, surgical treatment of rare diseases of the knee joint synovium. In diffuse forms of pigmented villous nodular synovitis, it is advisable to continue comparative studies of various surgical methods of capsular sinovectomy (open and arthroscopic techniques).

Enhanced osteochondral regeneration with a 3D-Printed biomimetic scaffold featuring a calcified interfacial layer

The integrative regeneration of both articular cartilage and subchondral bone remains an unmet clinical need due to the difficulties of mimicking spatial complexity in native osteochondral tissues for artificial implants. Layer-by-layer fabrication strategies, such as 3D printing, have emerged as a promising technology replicating the stratified zonal architecture and varying microstructures and mechanical properties. However, the dynamic and circulating physiological environments, such as mass transportation or cell migration, usually distort the pre-confined biological properties in the layered implants, leading to undistinguished spatial variations and subsequently inefficient regenerations. This study introduced a biomimetic calcified interfacial layer into the scaffold as a compact barrier between a cartilage layer and a subchondral bone layer to facilitate osteogenic–chondrogenic repair. The calcified interfacial layer consisting of compact polycaprolactone (PCL), nano-hydroxyapatite, and tasquinimod (TA) can physically and biologically separate the cartilage layer (TA-mixed, chondrocytes-load gelatin methacrylate) from the subchondral bond layer (porous PCL). This introduction preserved the as-designed independent biological environment in each layer for both cartilage and bone regeneration, successfully inhibiting vascular invasion into the cartilage layer and preventing hyaluronic cartilage calcification owing to devascularization of TA. The improved integrative regeneration of cartilage and subchondral bone was validated through gross examination, micro-computed tomography (micro-CT), and histological and immunohistochemical analyses based on an in vivo rat model. Moreover, gene and protein expression studies identified a key role of Caveolin (CAV-1) in promoting angiogenesis through the Wnt/β-catenin pathway and indicated that TA in the calcified layer blocked angiogenesis by inhibiting CAV-1.

A Finite Element Model to Investigate the Stability of Osteochondral Grafts Within a Human Tibiofemoral Joint

Osteochondral grafting has demonstrated positive outcomes for treating articular cartilage defects by replacing the damaged region with a cylindrical graft consisting of bone with a layer of cartilage. However, factors that cause graft subsidence are not well understood. The aim of this study was to develop finite element (FE) models of osteochondral grafts within a tibiofemoral joint, suitable for an investigation of parameters affecting graft stability. Cadaveric femurs were used to experimentally calibrate the bone properties and graft-bone frictional forces for use in corresponding image-based FE models, generated from µCT scan data. Effects of cartilage defects and osteochondral graft repair were measured by examining contact pressure changes using further in vitro tests. Here, six defects were created in the femoral condyles, which were subsequently treated with osteochondral autografts or metal pins. Matching image-based FE models were created, and the contact patches were compared. The bone material properties and graft-bone frictional forces were successfully calibrated from the initial tests with good resulting levels of agreement (CCC = 0.87). The tibiofemoral joint experiment provided a range of cases that were accurately described in the resultant pressure maps and were well represented in the FE models. Cartilage defects and repair quality were experimentally measurable with good agreement in the FE model pressure maps. Model confidence was built through extensive validation and sensitivity testing. It was found that specimen-specific properties were required to accurately represent graft behaviour. The final models produced are suitable for a range of parametric testing to investigate immediate graft stability.

Evaluation of rabbit adipose derived stem cells fate in perfused multilayered silk fibroin composite scaffold for Osteochondral repair.

Development of osteochondral tissue engineering approaches using scaffolds seeded with stem cells in association with mechanical stimulations has been recently considered as a promising technique for the repair of this tissue. In this study, an integrated and biomimetic trilayered silk fibroin (SF) scaffold containing SF nanofibers in each layer was fabricated. The osteogenesis and chondrogenesis of stem cells seeded on the fabricated scaffolds were investigated under a perfusion flow. 3-Dimethylthiazol-2,5-diphenyltetrazolium bromide assay showed that the perfusion flow significantly enhanced cell viability and proliferation. Analysis of gene expression by stem cells revealed that perfusion flow had significantly upregulated the expression of osteogenic and chondrogenic genes in the bone and cartilage layers and downregulated the hypertrophic gene expression in the intermediate layer of the scaffold. In conclusion, applying flow perfusion on the prepared integrated trilayered SF-based scaffold can support osteogenic and chondrogenic differentiation for repairing osteochondral defects.

Effects of tetrahedral DNA nanostructures on the treatment of osteoporosis.

Osteoporosis (OP) is a common disease characterized by bone loss and bone tissue microstructure degradation. Drug treatment is a common clinical treatment that aims to increase bone mass and bone density. Tetrahedral DNA nanostructures (TDNs) are three-dimensional tetrahedral frames formed by folding four single-stranded DNA molecules, which have good biological safety and can promote bone regeneration. In this study, a mouse model of OP was established by ovariectomy (OVX) and TDN was injected into the tail vein for 8 weeks. We found that ovariectomized mice could simulate some physiological changes in OP. After treatment with TDNs, some of this destruction in mice was significantly improved, including an increase in the bone volume fraction (BV/TV) and bone trabecular number (Tb. N), decrease in bone separation (Tb. SP), reduction in the damage to the mouse cartilage layer, reduction in osteoclast lacunae in bone trabecula, and reduction in the damage to the bone dense part. We also found that the expression of ALP, β-Catenin, Runx2, Osterix, and bone morphogenetic protein (BMP)2 significantly decreased in OVX mice but increased after TDN treatment. Therefore, this study suggests that TDNs may regulate the Wnt/β-Catenin and BMP signalling pathways to improve the levels of some specific markers of osteogenic differentiation, such as Runx2, ALP, and Osterix, to promote osteogenesis, thus showing a therapeutic effect on OP mice.

Arthrological reconstructions of the pterosaur neck and their implications for the cervical position at rest.

The lack of any pterosaur living descendants creates gaps in the knowledge of the biology of this group, including its cervical biomechanics, which makes it difficult to understand their posture and life habits. To mitigate part of this issue, we reconstructed the cervical osteology and arthrology of three pterosaurs, allowing us to make inferences about the position of the neck of these animals at rest. We used scans of three-dimensionally preserved cervical series of Anhanguera piscator, Azhdarcho lancicollis and Rhamphorhynchus muensteri for the reconstructions, thus representing different lineages. For the recognition of ligaments, joint cartilages, and levels of overlapping of the zygapophyses, we applied the Extant Phylogenetic Bracket method, based on various extant birds and on Caiman latirostris. We inferred that pterosaur intervertebral joints were probably covered by a thin layer of synovial cartilage whose thickness varied along the neck, being thicker in the posterior region. Ignoring this cartilage can affect reconstructions. According to the vertebral angulation, their neck was slightly sinuous when in rest position. Our analyses also indicate that pterosaurs had segmented and supra-segmented articular cervical ligaments, which could confer stabilization, execute passive forces on the neck and store elastic energy.

Influence of leg axis alignment on MRI T2* mapping of the knee in young professional soccer players

BackgroundInvestigation of the association between leg axis alignment and biochemical MRI in young professional soccer players in order to identify a potential influence of the leg axis on cartilage regions at risk.MethodsSixteen professional soccer players (21 ± 3 years) underwent static and dynamic leg axis analysis via radiation free DIERS formetric 4 D as well as 3-T MRI examination of both knees. Quantitative T2* mapping of the knee cartilage was performed and T2* values were evaluated as 144 regions of interest. Subgroup analysis was performed in players with severe varus alignment (> 6°).ResultsAnalysis of the leg axis geometry revealed a mean static alignment of 6.6° ± 2.5 varus and a mean dynamic alignment of 5.1° ± 2.6 varus. Quantitative T2* mapping showed significantly increased T2* values in the superficial cartilage layer compared to the deeper region (p < 0.001) as well as a significant increase in relaxation times in the femoral cartilage from anterior to intermediate to posterior (p < 0.001). Combination of both methods revealed a significant correlation for the degree of varus alignment and the femoral, posterior, deep region of the medial knee compartment (r = 0.4; p = 0.03). If severe varus alignment was present this region showed a significant increase in relaxation time compared to players with a less pronounced leg axis deviation (p = 0.003).ConclusionThis study demonstrates that varus alignment in young soccer players is associated with elevated T2* relaxation times in the deep cartilage layer of the medial, posterior, femoral compartment and might therefore be a contributing factor in the early pathogenesis of manifest cartilage lesions. Therefore, these findings should be considered in the development of preventive training programs.

Confirmation of pain-related neuromodulation mechanism of Bushen Zhuangjin Decoction on knee osteoarthritis

Ethnopharmacological relevanceBushen Zhuangjin Decoction (BZD) are an herbal compound commonly used to treat osteoarthritis (OA) in China. Aim of the studyThis study aimed to verify the mechanism of Bushen Zhuangjin Decoction in relieving the pain of knee osteoarthritis. Materials and methodsNetwork pharmacology evaluation was used to discover the potential targets of BZD to relieve pain in KOA. The therapeutic effects of BZD treatment on KOA pain using histomorphology, behavioral assessments, suspension chip analysis, and ultra-high performance liquid chromatography/tandem mass spectrometry (UHPLC-MS/MS) assays. The functional magnetic resonance imaging was used to explore the effects of BZD treatment on brain function associated to KOA. ResultsNetwork pharmacological analysis revealed the association between the analgesic effect of BZD on KOA and the pain signaling neurotransmitter 5-HT. Subsequently, we conducted experiments to verify the therapeutic effect of BZD on pain in KOA animal models. Behavioral tests demonstrated that the pain threshold of knee osteoarthritis rats decreased in PWT and PWL, but BZD was able to increase the pain threshold. Histopathological staining indicated thinning of the cartilage layer and sparse trabeculae in the subchondral bone. Suspension chip analysis revealed a significant increase in pro-inflammatory factors of IL-1α, IL-5, IL-12, IL-17A, RANTES, TNF-α and M-CSF in KOA, along with a significant decrease in anti-inflammatory factor of IL-13. However, BZD treatment decreased the expression of pro-inflammatory factors and increased the content of anti-inflammatory factor. UHPLC-MS/MS analysis showed a significant decrease in the serum levels of GABA, E, GSH, Kyn, Met, and VMA in KOA, which were significantly increased by BZD. Conversely, the serum levels of TrpA, TyrA, Spd, and BALa were significantly increased in KOA and significantly decreased by BZD. ELISA and Western blot analysis showed increased expression of subchondral bone pain-related neuropeptides SP, CGRP, TH, NPY, VEGFA, 5-HT3 in KOA, which were decreased in BZD. Functional magnetic resonance imaging demonstrated that BZD exerts its therapeutic effect on KOA by modulating the activity and functional connections of the cortex, hypothalamus, and hippocampus. ConclusionsThis study confirmed the significant role of pain-related neuromodulation mechanisms in the analgesic therapy of BZD and provides a theoretical foundation for using BZD as a traditional Chinese medical treatment for KOA pain.

Three‐dimensional printed biomimetic multilayer scaffolds coordinated with sleep‐related small extracellular vesicles: A strategy for extracellular matrix homeostasis and macrophage polarization to enhance osteochondral regeneration

AbstractCartilage defects resulting from injury or degeneration are a common clinical problem, and due to its avascular nature, articular cartilage has poor self‐healing capacity. Three‐dimensional (3D) bioprinting has attracted great attention in tissue engineering. Melatonin (MT), a hormone mainly secreted at night, plays an important role in tissue repair. Small extracellular vesicles (sEV) are considered ideal drug delivery vehicles and MT‐sEV (sleep‐related sEV) have the potential ability to promote cartilage regeneration. Here, biomimetic multilayer scaffolds were fabricated using 3D bioprinting. A double network hydrogel, composed of methacrylated hyaluronic acid and gelatin methacryloyl (HG), was prepared. MT‐sEV and HG hydrogel were used to create a cartilage layer. A bone layer was formed using poly(ε‐caprolactone) and hydroxyapatite ultralong nanowires. Additionally, two bioinks were alternately printed at the interface layer. The results of RNA sequencing revealed the potential regulatory mechanisms. MT‐sEV showed promotional effects on cell migration, proliferation, chondrogenic differentiation, and extracellular matrix (ECM) deposition. Moreover, MT‐sEV altered macrophage polarization and regulated the expression of inflammatory cytokines. In vivo experiments demonstrated that the biomimetic multilayer scaffolds promoted cartilage regeneration. These experiments demonstrated the ability of MT‐sEV to regulate the immune microenvironment and promote the secretion of ECM, providing a promising strategy for cartilage regeneration.

Composite Spheroid-Laden Bilayer Hydrogel for Engineering Three-Dimensional Osteochondral Tissue.

A combination of hydrogels and stem cell spheroids has been used to engineer three-dimensional (3D) osteochondral tissue, but precise zonal control directing cell fate within the hydrogel remains a challenge. In this study, we developed a composite spheroid-laden bilayer hydrogel to imitate osteochondral tissue by spatially controlled differentiation of human adipose-derived stem cells. Meticulous optimization of the spheroid-size and mechanical strength of gelatin methacryloyl (GelMA) hydrogel enables the cells to homogeneously sprout within the hydrogel. Moreover, fibers immobilizing transforming growth factor beta-1 (TGF-β1) or bone morphogenetic protein-2 (BMP-2) were incorporated within the spheroids, which induced chondrogenic or osteogenic differentiation of cells in general media, respectively. The spheroids-filled GelMA solution was crosslinked to create the bilayer hydrogel, which demonstrated a strong interfacial adhesion between the two layers. The cell sprouting enhanced the adhesion of each hydrogel, demonstrated by increase in tensile strength from 4.8 ± 0.4 to 6.9 ± 1.2 MPa after 14 days of culture. Importantly, the spatially confined delivery of BMP-2 within the spheroids increased mineral deposition and more than threefold enhanced osteogenic genes of cells in the bone layer while the cells induced by TGF-β1 signals were apparently differentiated into chondrocytes within the cartilage layer. The results suggest that our composite spheroid-laden hydrogel could be used for the biofabrication of osteochondral tissue, which can be applied to engineer other complex tissues by delivery of appropriate biomolecules.

Middle Ear Ossicular Joint Changes in Type 2 Diabetes Mellitus: A Histopathological Study.

Although previous research has indicated inner ear changes in diabetes mellitus (DM) patients, no prior study has explored the middle ear, particularly the ossicles and their joints, in DM patients. This study aimed to investigate whether type 2 DM is associated with middle ear changes, specifically affecting the ossicular chain and joints. This study included 47 ears from 25 patients with DM (male = 13, female = 12, age: 51.0 ± 20.5) and age- and sex-matched controls (male = 10, female = 10, age: 54.8 ± 15.9) (sex; p = 1.000, Age; p = 0.991). Otopathological evaluations of the auditory ossicles and incudomalleolar joint (IMJ) were performed using light microscopy. In the IMJ of DM cases, malleus hyalinized cartilage (Malleus hC) and incus hyalinized cartilage (Incus hC) were significantly increased compared with control cases (Malleus hC; DM, 34.17 ± 9.71 μm vs. control 21.96 ± 4.16 μm, p < 0.001) (Incus hC; DM 35.11 ± 10.12 μm vs. control 22.42 ± 4.368 μm, p < 0.001). In addition, bone-line distance was significantly longer than in DM cases than control cases (DM 266.72 ± 59.11 μm vs. control 239.81 ± 35.56 μm p = 0.040). On the other hand, joint discus distance was longer in the control group than in DM cases (DM 96.84 ± 36.80 μm vs. Control 113.63 ± 23.81 μm, p = 0.001). This study reveals a notable increase in the hyalinized cartilage layer and bone-line distance accompanied by reducing joint discus distance within the IMJ in DM cases. These findings suggest that DM may influence microjoints, such as the IMJ, and potentially impact auditory function. N/A Laryngoscope, 134:2871-2878, 2024.

TARGETING HYPERTROPHIC CHONDROCYTES IN OSTEOARTHRITIS: THE ROLE OF INFLAMMATION AND THE USE OF A MULTI-MODEL APPROACH IN SEARCH OF PHARMACOLOGICAL TREATMENTS

In osteoarthritis, chondrocytes acquire a hypertrophic phenotype that contributes to matrix degradation. Inflammation is proposed as trigger for the shift to a hypertrophic phenotype. Using in vitro culture of human chondrocytes and cartilage explants we could not find evidence for a role of inflammatory signalling activation. We found, however, that tissue repair macrophages may contribute to the onset of hypertrophy (doi: 10.1177/19476035211021907) Intra-articularly injected triamcinolone acetonide to inhibit inflammation in a murine model of collagenase-induced osteoarthritis, increased synovial macrophage numbers and osteophytosis, confirming the role of macrophages in chondrocyte hypertrophy occurring in osteophyte formation (doi: 10.1111/bph.15780).In search of targets to inhibit chondrocyte hypertrophy, we combined existing microarray data of different cartilage layers of murine growth plate and murine articular cartilage after induction of collagenase-induced osteoarthritis. We identified common differentially expressed genes and selected those known to be associated to inflammation. This revealed EPHA2, a tyrosine kinase receptor, as a new target. Using in silico, in vitro and in vivo models we demonstrated that inhibition of EPHA2 might be a promising treatment for osteoarthritis.Recently, single cell RNA-seq. has revealed detailed information about different populations of chondrocytes in articular cartilage during osteoarthritis. We re-analysed a published scRNA-seq data set of healthy and osteoarthritic cartilage to obtain the differentially expressed genes in the population of hypertrophic chondrocytes compared to the other chondrocytes, applied pathway analyses and then used drug databases to search for upstream inhibitors of these pathways. This drug repurposing approach led to the selection of 6 drugs that were screened and tested using several in vitro models with human chondrocytes and cartilage explants.In this lecture I will present this sequence of studies to highlight different approaches and models that can be used in the quest for a disease modifying drug for osteoarthritis.

THE POTENTIAL OF MAGNESIUM-BASED MICRO CYLINDER FOR CARTILAGE AND BONE REGENERATION UTILIZING AN IN VITRO OSTEOARTHRITIS MODEL

Osteoarthritis (OA) is an inflammatory disease affecting the complete synovial joint including the cartilage layer and the subchondral bone plate. Due to the multifactorial causes and the not yet completely resolved molecular mechanisms, it lacks a gold standard treatment to mitigate OA. Hence, biomaterials capable of delaying or preventing OA are a promising alternative or supplement to antiphlogistic and surgical interventions. Magnesium (Mg) and its alloys are among the promising biomaterials with osteoinductive effects. This work investigated the impact of Mg micro cylinders (length ≈of 1.0 mm and width of 0.5 mm) in vitro, in favoring joint regeneration together with preventing OA progression. Therefore, a mesenchymal stem cell line (SCP-1) was applied in order to assess the compatibility of the degradable material. Furthermore, an in vitro OA model utilizing SCP-1 cells based on the supplementation of the cytokines; IL-1β, TNF-α was established and disclosed the capability of Mg microparticles in differentiating SCP-1 cells into chondrogenic and osteogenic lineages proven through extracellular matrix staining and gene marker analysis. A concentration above 10 mM revealed a reduction in the cell viability by 50 %. An increase in the expression of collagens especially and proteoglycans (COL2A1, Aggrecan) as extracellular matrix proteins as well as an increase in osteogenic marker (ALP, BMP2) favoring the mineralization process were observed. The inflammatory condition reduced the viability and productivity of the applied stem cell line. However, the application of Mg microparticles induced a cell recovery and reduction of inflammation marker such as MMP1 and IL6. The cytocompatible and the ability of Mg microparticles in supporting bone and cartilage repair mechanisms in vitro even under inflammatory conditions make biodegradable Mg microparticles a suitable implant material to treat OA therapy.Acknowledgements: This project OAMag was funded by the German Research Foundation (project number 404534760). The author thank Dr. Björn Wiese (hereon) for the production of Mg based material and Prof. Böcker (MUM Musculoskeletal University Center Munich) for the provision of SCP-1 cell line.

MULTILAYER DUAL-POROSITY 3D-PRINTED SCAFFOLDS TO RECREATE THE ANISOTROPIC MICROENVIRONMENT OF THE CARTILAGE

Articular cartilage is a multi-zonal tissue that coats the epiphysis of long bones and avoids its wear during motion. An unusual friction could micro-fracture this connective membrane and progress into an osteochondral defect (OD), where the affected cartilage suffers inflammation, fibrillation, and forfeiture of its anisotropic structure.Clinical treatment for ODs has been focused on micro-fracture techniques, where the defect area is removed and small incisions are performed in the subchondral bone, which allows the exudation of mesenchymal stem cells (hMSCs) to the abraded zone. However, hMSCs represent less than 0.01% of the total cell population and are not able to self-organise coherently, so the treatments fail in the long term. To select, support and steer hMSCs from the bone marrow into a specific differentiation stage, and recreate the cartilage anisotropic microenvironment, multilayer dual-porosity 3D-printed scaffolds were developed.Dual-porosity scaffolds were printed using prepared inks, containing specific ratios of poly-(d,l)lactide-co-caprolactone copolymer and gelatine microspheres of different diameters, which acted as sacrificial micro-pore templates and were leached after printing. The cell adhesion capability was investigated showing an increased cell number in dual-porosity scaffolds as compared to non-porous ones. To mimic the stiffness of the three cartilage zones, several patterns were designed, printed, and checked by dynamic-mechanical analysis under compression at 37 ºC. Three patterns with specific formulations were chosen as candidates to recreate the mechanical properties of the cartilage layers. Differentiation studies in the selected scaffolds showed the formation of mature cartilage by gene expression, protein deposition and biomolecular analysis. Given the obtained results, designed scaffolds were able to guide hMSC behaviour.In conclusion, biocompatible, multilayer and dual-porosity scaffolds with cell entrapment capability were manufactured. These anisotropic scaffolds were able to recreate the physical microenvironment of the natural cartilage, which in turn stimulated cell differentiation and the formation of mature cartilage.Acknowledgments: This work was supported by the EMAKIKER grant.

Comparison of Arthroscopic Microfracture and Retrograde Subchondral Drilling in the Treatment of Osteochondral Lesions of Talus.

In our study, it was aimed to evaluate the efficacy of microfracture and retrograde subchondral drilling on clinical outcomes in patients who underwent ankle arthroscopy due to osteochondral lesion of medial talus. Twenty eight patients with osteochondral lesion of talus less than 1.5 cm2 treated with ankle arthroscopy were evaluated retrospectively. Microfracture was performed in 16 patients and retrograde subchondral drilling was performed in 12 patients that there was no loss of integrity in the cartilage layer or cartilage layer is partially preserved during ankle arthroscopy. Postoperative evaluations of the patients were done with Foot and Ankle Ability Measure (FAAM) and results of both group were compared statistically. The mean activities of daily living scale was 93.4±3.2 and sportive activity scale was 90.1±5.7 in the retrograde drilling group. In the micro-fracture group, mean activities of daily living scale was 93.8±4.1 and mean sportive activity scale was 88.9±9.5. No significant difference was found as a result of statistical comparisons of both groups results. It has been determined that the size and preservation of the integrity of talar osteochondral lesions are important factors on clinical results. Results of arthroscopic debridement, microfracture and drilization are not good in lesions larger than 1.5 cm2 and lesions with impaired integrity. In our study, depending on the general literature, osteochondral lesions in talus were less than 1.5 cm2 in patients who underwent arthroscopic micro fracture and retrograde drilling. Both microfracture and retrograde subchondral drilling are effective treatment methods with good clinical results for talar osteochondral lesions less than 1,5cm2 . Retrograde subchondral drilling can be an alternative treatment method with the reliability of clinical results in patients with no loss of the integrity of the cartilage layer or cartilage layer is partially preserved. talus, osteochondral lesion, microfracture, subchondral drilling.

Histological differences in cartilage layer growth at various tendon and ligament insertions in rabbits.

Objectives: Histological differences in cartilage layer growth in Achilles tendon (AT), quadriceps tendon (QT), patellar tendon (PT), and anterior cruciate ligament (ACL) insertion are unclear. Therefore, this study aimed to investigate the differences in cartilage layer growth in AT, QT, PT, and ACL insertions. Materials and Methods: Forty-eight male Japanese white rabbits were used. Six animals were euthanized at different stages (day 1 and 1, 2, 4, 6, 8, 12, and 24 weeks). Safranin O-stained glycosaminoglycan (GAG) production area, chondrocyte count, and insertion width were investigated. Results: A two-way analysis of variance (ANOVA) revealed a significant difference in the main effects of time and insertion for all parameters. In addition, the time × insertion interaction was significant. Multiple comparisons showed a significant difference between the ACL insertion and all other variables; however, the GAG production area was not significantly different for the QT, PT, and AT insertions. AT insertions were significantly different from all other groups; however, the number of chondrocytes and insertion width were not significantly different for ACL, QT, and PT insertions. Conclusion: Cartilage layer growth differed between the AT, QT, PT, and ACL insertions. The differences between the insertions may also be due to the differences in their structures, locations, and mechanical environments.

Osteochondral tissue engineering in translational practice: histological assessments and scoring systems.

Osteochondral lesions are common pathological alterations in synovial joints. Different techniques have been designed to achieve osteochondral repair, and tissue-engineered osteochondral grafts have shown the most promise. Histological assessments and related scoring systems are crucial for evaluating the quality of regenerated tissue, and the interpretation and comparison of various repair techniques require the establishment of a reliable and widely accepted histological method. To date, there is still no consensus on the type of histological assessment and scoring system that should be used for osteochondral repair. In this review, we summarize common osteochondral staining methods, discuss the criteria regarding high-quality histological images, and assess the current histological scoring systems for osteochondral regeneration. Safranin O/Fast green is the most widely used staining method for the cartilage layer, whereas Gomori and Van Gieson staining detect new bone formation. We suggest including the graft-host interface and more sections together with the basic histological information for images. An ideal scoring system should analyze both the cartilage and bone regions, especially for the subchondral bone plate. Furthermore, histological assessments should be performed over a longer period of time to minimize discrepancies caused by defect size and animal species.