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As an indispensable component of the extracellular matrix, perlecan (Pln) plays an essential role in cartilaginous tissue function. Although there exist studies suggesting that Pln expressed by cartilaginous tissues is critical for chondrogenesis, few papers have discussed the potential impact Pln may have on cartilage regeneration. In this review, we delineate Pln structure, biomechanical properties, and interactive ligands-which together contribute to the effect Pln has on cartilaginous tissue development. We also review how the signaling pathways of Pln affect cartilage development and scrutinize the potential application of Pln to divisions of cartilage regeneration, spanning vascularization, stem cell differentiation, and biomaterial improvement. The aim of this review is to deepen our understanding of the spatial and temporal interactions that occur between Pln and cartilaginous tissue and ultimately apply Pln in scaffold design to improve cell-based cartilage engineering and regeneration. STATEMENT OF SIGNIFICANCE: As a key component of the basement membrane, Pln plays a critical role in tissue development and repair. Recent findings suggest that Pln existing in the pericellular matrix surrounding mature chondrocytes is actively involved in cartilage regeneration and functionality. We propose that Pln is essential to developing an in vitro matrix niche within biological scaffolds for cartilage tissue engineering.

Relapsing polychondritis (RP) is a rare multisystem disorder characterized by inflammation and destruction of cartilaginous tissue. Atypical initial symptoms and signs may cause difficulties in the diagnosis of RP for pediatricians. In this article, we report a 9-year-old male who presented with stridor and hoarseness indicating upper respiratory tract involvement in the setting of chronic polyarthralgia. Flexible bronchoscopy revealed subglottic stenosis. Increasing awareness of early respiratory manifestations of RP will prevent life-threatening complications.

A major shortcoming in cartilage tissue engineering is the low biosynthetic response of chondrocytes. While different strategies have been investigated, a novel approach may be to control nutrient metabolism. Although known for their anaerobic metabolism, chondrocytes are more synthetically active under conditions that elicit mixed aerobic-anaerobic metabolism. Here, we postulate this metabolic switch induces HIF-1α signaling resulting in improved growth. Transition to different metabolic states can result in pooling of metabolites, several of which can stabilize HIF-1α by interfering with PHD2. Chondrocytes cultured under increased media availability accelerated tissue deposition with the greatest effect occurring at 2 mL/106 cells. Under higher media availability, metabolism switched from anaerobic to mixed aerobic-anaerobic. Around this transition, maximal changes in PHD2 activity, HIF-1α expression, and HIF-1 target gene expression were observed. Loss-of-function studies using YC-1 confirmed the involvement of HIF-1. Lastly, targeted metabolomic studies revealed that intracellular lactate and succinate correlated with PHD2 activity. This study demonstrates that cartilaginous tissue formation can be regulated by nutrient metabolism and that this response is mediated through changes in HIF-1α signalling. By harnessing this newly identified metabolic switch, engineered cartilage implants may be developed without the need for sophisticated methods which could aid translation to the clinic. This article is protected by copyright. All rights reserved.

Pankiv M. V., Paltov Ye. V., Masna Z. Z., Chelpanova I. V., Kovalska M. Ye. Structural features of the intervertebral disc in rats. Journal of Education, Health and Sport. 2021;11(07): 68-78. eISSN 2391-8306. DOI http://dx.doi.org/10.12775/JEHS.2021.11.07.006 https://apcz.umk.pl/czasopisma/index.php/JEHS/article/view/JEHS.2021.11.07.006 https://zenodo.org/record/5093282 The journal has had 5 points in Ministry of Science and Higher Education parametric evaluation. § 8. 2) and § 12. 1. 2) 22.02.2019. © The Authors 2021; This article is published with open access at Licensee Open Journal Systems of Nicolaus Copernicus University in Torun, Poland Open Access. This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author (s) and source are credited. This is an open access article licensed under the terms of the Creative Commons Attribution Non commercial license Share alike. (http://creativecommons.org/licenses/by-nc-sa/4.0/) which permits unrestricted, non commercial use, distribution and reproduction in any medium, provided the work is properly cited. The authors declare that there is no conflict of interests regarding the publication of this paper. Received: 14.06.2021. Revised: 25.06.2021. Accepted: 12.07.2021. UDK: 611.721.1.018.3.019 Structural features of the intervertebral disc in rats M. V. Pankiv, Ye. V. Paltov, Z. Z. Masna, I. V. Chelpanova, M. Ye. Kovalska Danylo Halytsky Lviv National Medical University, Lviv, Ukraine M.V. Pankiv, Ph. D-student at the Department of Normal Anatomy, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine; https://orcid.org/0000-0002-3714-2577 Ye.V. Paltov, Ph.D., Associate Professor, Head of the Laboratory of Electron Microscopy, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine; https://orcid.org/0000-0002-2622-4753 Z.Z. Masna, MD, Professor, Head of the Department of Operative Surgery and Topographical Anatomy, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine; https://orcid.org/0000-0003-2057-7061 I.V. Chelpanova, Ph.D., Associate Professor, Department of Histology, Cytology and Embryology, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine https://orcid.org/ 0000-0001-5215-814X M. Ye. Kovalska Ph.D., Assistant of the department of pathological physiology, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine; https://orcid.org/0000-0002-6232-3951 Abstract The aim of our work was to study the structural organization of the components of vertebral disc of rats on the microscopic level in the normal state. This goal was achieved by means of microscopic method of visualization of cellular components of the intervertebral disc of rats. Histological preparations were made in accordance with the conventional method of using the dye hematoxylin, eosin and azan according to the method of Heidenhain, PAS reaction according to MacManus and Alcian blue according to Steedman. As a result of the microstructural study, it was ascertained that the rat’s intervertebral disc normally consists of a gelatinous (pulpal) nucleus and a fibrous ring. It is separated from the vertebral bodies by a thin layer of cartilaginous tissue of the locking plates and hyaline cartilage, which covers the bodies of adjacent vertebrae. The data that rats do not have locking plates can be rarely found in the literature. The vessels verging the intervertebral disc radially -in front, back and from the sides, branch out from the vessels of vertebral periosteum. In the cartilaginous tissue of intervertebral disc, vessels are not visualized. Key words: intervertebral disc; structural components; norm; rat.

3D bioprinting has emerged as a promising technology in the field of tissue engineering and regenerative medicine due to its ability to create anatomically complex tissue substitutes. However, it still remains challenging to develop bioactive bioinks that provide appropriate and permissive environments to instruct and guide the regenerative process in vitro and in vivo. In this study alginate sulfate, a sulfated glycosaminoglycan (sGAG) mimic, was used to functionalize an alginate-gelatin methacryloyl (GelMA) interpenetrating network (IPN) bioink to enable the bioprinting of cartilaginous tissues. The inclusion of alginate sulfate had a limited influence on the viscosity, shear-thinning and thixotropic properties of the IPN bioink, enabling high-fidelity bioprinting and supporting mesenchymal stem cell (MSC) viability post-printing. The stiffness of printed IPN constructs greatly exceeded that achieved by printing alginate or GelMA alone, while maintaining resilience and toughness. Furthermore, given the high affinity of alginate sulfate to heparin-binding growth factors, the sulfated IPN bioink supported the sustained release of transforming growth factor-β3 (TGF-β3), providing an environment that supported robust chondrogenesis in vitro, with little evidence of hypertrophy or mineralization over extended culture periods. Such bioprinted constructs also supported chondrogenesis in vivo, with the controlled release of TGF-β3 promoting significantly higher levels of cartilage-specific extracellular matrix deposition. Altogether, these results demonstrate the potential of bioprinting sulfated bioinks as part of a 'single-stage' or 'point-of-care' strategy for regenerating cartilaginous tissues. STATEMENT OF SIGNIFICANCE: This study highlights the potential of using sulfated interpenetrating network (IPN) bioink to support the regeneration of phenotypically stable articular cartilage. Construction of interpenetrating networks in the bioink enables unique high-fidelity bioprinting and provides synergistic increases in mechanical properties. The presence of alginate sulfate enables the capacity of high affinity-binding of TGF-β3, which promoted robust chondrogenesis in vitro and in vivo.

A 13-year-old castrated male Miniature Dachshund exhibited oedema of the face and cervical region. Clinical examination, including computed tomography, revealed a mass on the right atrial wall. Histopathological examination of the mass revealed proliferation of spindle cells, which formed intersecting bundles and abundant proteoglycan material. Multiple islands of hyaline cartilage were also observed within foci of proliferated spindle cells. Although the proliferated spindle cells replaced the myocardial parenchyma, there was no evidence of malignancy. The spindle cells were immunopositive for vimentin and α-smooth muscle actin, while chondrocytes were immunopositive for vimentin and S100. Neither the spindle cells nor the chondrocytes were immunolabelled for cytokeratin AE1/AE3, desmin, factor VIII, melan A, p63 or Ki-67. These findings indicate that the lesion represented myocardial fibrosis with cartilaginous tissue formation.

We report the peculiar case of a parosteal osteosarcoma arising beneath the periosteum in a 12-year-old boy. He complained of difficulty in left knee flexion. Plain radiography showed a uniformly dense mineralized mass in the bone cortex and parosteal ossified nodules at the metaphysis and diaphysis of the left distal femur. Periosteal reaction was not evident. Uniquely, plain radiography had a smooth outline and revealed gradually thickening mass toward the center. Histologically, the tumor showed a proliferation of spindle-shaped cells with parallel-oriented dense bone trabeculae and hyaline cartilaginous tissue disclosing mild atypia. The periosteum was inverted along the polypoid mass, but there was no periosteum at the top. Immunohistochemically, the spindle cells, including those at the top of the polypoid mass, and cartilaginous cells were positive for MDM2 and CDK4. MDM2 gene amplification was detected in these cells by fluorescence in situ hybridization. Despite the peculiar feature of plain radiography, the lesion was diagnosed as parosteal osteosarcoma. This case report presents a case of parosteal osteosarcoma arising beneath the periosteum, although it is postulated to arise in the outer layer of the periosteum. The unique radiographic findings in this case suggest an association of parosteal osteosarcoma with vigorous bone growth before closure of the growth plate.

The strain-generated potential (SGP) is a well-established mechanism in cartilaginous tissues whereby mechanical forces generate electrical potentials. In articular cartilage (AC) and the intervertebral disc (IVD), studies on the SGP have focused on fluid- and ionic-driven effects, namely Donnan, diffusion and streaming potentials. However, recent evidence has indicated a direct coupling between strain and electrical potential. Piezoelectricity is one such mechanism whereby deformation of most biological structures, like collagen, can directly generate an electrical potential. In this review, the SGP in AC and the IVD will be revisited in light of piezoelectricity and mechanotransduction. While the evidence base for physiologically significant piezoelectric responses in tissue is lacking, difficulties in quantifying the physiological response and imperfect measurement techniques may have underestimated the property. Hindering our understanding of the SGP further, numerical models to-date have negated ferroelectric effects in the SGP and have utilised classic Donnan theory that, as evidence argues, may be oversimplified. Moreover, changes in the SGP with degeneration due to an altered extracellular matrix (ECM) indicate that the significance of ionic-driven mechanisms may diminish relative to the piezoelectric response. The SGP, and these mechanisms behind it, are finally discussed in relation to the cell response.

Intramyocardial cartilage has never been reported in the human heart before. In the literature, the only reported localizations of cartilage in the heart were in the central fibrous body and the valves. We report a case of an unusual presence of cartilage tissue within the myocardial wall of the left ventricle in a 10-year-old boy who died unexpectedly. This case presents an interesting, unusual and apparently asymptomatic sudden cardiac death related to a cartilaginous myocardial tumor. Conducting system disturbance secondary to the myocardial tumor is the probable cause of death. This case is relevant not only for its singularity and originality, but also for the diverse and controversial hypotheses related to the onset of cartilaginous tissue in the myocardial wall. Early detection of this tumor by modern thoracic imaging may have prevented a fatal unexpected outcome.

Biomechanical changes to the collagen fibrillar architecture in articular cartilage are believed to play a crucial role in enabling normal joint function. However, experimentally there is little quantitative knowledge about the structural response of the Type II collagen fibrils in cartilage to cyclic loading in situ, and the mechanisms that drive the ability of cartilage to withstand long-term repetitive loading. Here we utilize synchrotron small-angle X-ray scattering (SAXS) combined with in-situ cyclic loading of bovine articular cartilage explants to measure the fibrillar response in deep zone articular cartilage, in terms of orientation, fibrillar strain and inter-fibrillar variability in healthy cartilage and cartilage degraded by exposure to the pro-inflammatory cytokine IL-1β. We demonstrate that under repeated cyclic loading the fibrils reversibly change the width of the fibrillar orientation distribution whilst maintaining a largely consistent average direction of orientation. Specifically, the effect on the fibrillar network is a 3-dimensional conical orientation broadening around the normal to the joint surface, inferred by 3D reconstruction of X-ray scattering peak intensity distributions from the 2D pattern. Further, at the intrafibrillar level, this effect is coupled with reversible reduction in fibrillar pre-strain under compression, alongside increase in the variability of fibrillar pre-strain. In IL-1β degraded cartilage, the collagen rearrangement under cyclic loading is disrupted and associated with reduced tissue stiffness. These finding have implications as to how changes in local collagen nanomechanics might drive disease progression or vice versa in conditions such as osteoarthritis and provides a pathway to a mechanistic understanding of such diseases. Statement of significance Structural deterioration in biomechanically loaded musculoskeletal organs, e.g., joint osteoarthritis and back pain, are linked to breakdown and changes in their collagen-rich cartilaginous tissue matrix. A critical component enabling cartilage biomechanics is the ultrastructural collagen fibrillar network in cartilage. However, experimental probes of the dynamic structural response of cartilage collagen to biomechanical loads are limited. Here, we use X-ray scattering during cyclic loading (as during walking) on joint tissue to show that cartilage fibrils resist loading by a reversible, three-dimensional orientation broadening and disordering mechanism at the molecular level, and that inflammation reduces this functionality. Our results will help understand how changes to small-scale tissue mechanisms are linked to ageing and osteoarthritic progression, and development of biomaterials for joint replacements., Graphical abstract Image, graphical abstract

Introduction/purpose Osteochondral lesion of the talus (OCLT) is defined as a defect on the articular surface of the talus with/without subchondral bone involvement. Several surgical techniques are described in literature to treat OCLT. Particulated Juvenile Cartilage Allograft Transplantation (PJCAT) is a new emerging technique that has the potential of restoring the native cartilage. However, available data on PJCAT in treating OCLT is confined to a small number of studies with heterogeneous population. The aim of this study is to systematically review the literature on clinical and radiological outcomes of PJCAT in treating OCLT. Methods A systematic search of Medline database was performed using Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. Number of patients and demographic data included age, gender and body mass index (BMI) were extracted. The level of evidence of each included study was identified. When feasible, mechanism of injury, lesion size and average follow up were recorded. The American Orthopedic Foot and Ankle Society Score (AOFAS), and Foot and Ankle Outcome Score (FAOS) were obtained to assess the functional outcomes. Magnetic Resonance Observation of Cartilage Repair Tissue (MOCART) score was selected to evaluate the radiological outcomes. Results A total of 10 studies were eligible for this review involving 132 patients. Postoperative AOFAS scores were available for 44 patients who underwent PJCAT with an average of 86.14 at 25.5 months follow up. FAOS scores have been reported in 3 studies for 81 patients. The average preoperative score was 47.35 which has been improved to 62.88 at follow up of 23.6 months. MOCART was evaluated in 42 feet after the mean follow up of 18.2 months. The repair tissue was characterized by variable features; however, certain peculiarities were observed with higher frequency including infill hypertrophy, incomplete border zone integration, deep surface disruption, structure inhomogeneity, patches of hyperintense signals, damage of subchondral lamina and subchondral bone, adhesions and absence of effusion. Conclusion PJCAT seems to be a promising modality of treatment for OLT in terms of functional outcomes. Fair attempts of defect filling can be confirmed with MRI. Nevertheless, heterogeneous picture of regenerate cartilaginous tissue and lack of repair in subchondral bone and subchondral lamina are not in favor with claims of full restoration of lost normal hyaline articular cartilage.

Injuries and degenerative changes of tendons are common damages of the musculoskeletal system. Due to its hypovascular character the tendon has a limited natural ability to recover. For typical surgical treatment, the tendon integrity is restored, but in most cases, there occurs formation of the connective tissue scar resulting in structural and mechanical functionality disruption. The insufficient effectiveness of traditional therapy methods requires the search for alternative ways to restore damaged tendon tissues. This article discusses new effective methods for improving the treatment that base on the use of cellular technologies among which one of the main directions is mesenchymal stem cell application. Due to mesenchymal stem cells, there is a shift from pro-fibrotic and pro-inflammatory reactions of cells to pro-regenerative ones. Stem cells being multipotent and having among other things tenogenic potential are considered a promising material for repairing damaged tendons. The article also describes the sources of progenitor tendon cells including the tendon bundles and pericytes the main markers of which are Scx and Mkx that are proteins of the transcription factor superfamily, and Tnmd that is transmembrane glycoprotein.The growth factors that not only enhance the proliferative activity of mesenchymal stem cells but also promote in vitro tenogenic genes expression as well as the collagen Itype production what is necessary for tendon formation are considered. Along with growth factors, the morphogenetic protein BMP14 is presented, this protein increases themesenchymal stem cell proliferation and contributes directed tenogenic differentiation of these cells, suppressing their adipogenic and chondrogenic potentials.In recent years, mesenchymal stem cells have been used both separately and in combination with various growth factors and different three-dimensional structures providing the interaction with all of the cell types.The issues of the latest 3D-bioprinting technology allowing to make tissue-like structures for replacement damaged tissues and organs are discussed. 3D-bioprinting technology is known to allow acting exact spatio-temporal control of the distribution of cells, growth factors, small molecules, drugs and biologically active substances.

The results of experimental work on modeling deforming osteoarthrosis of the knee joint in small rodents are presented. The pathogenetic basis of this process is a violation of the blood supply and the distribution of physiological load in the joint. Variants of rigid and non-rigid limb immobilization systems are proposed. The results of morphofunctional assessment are given, indicating a violation of the anatomical and histological structure of the metaphysis, pineal gland and metaepiphysis of the femur and tibia. The leading role in the development of deforming osteoarthritis of the knee belongs to the violation of the physiological distribution of the dynamic load on the joint, and the most vulnerable to the progression of this process are subchondral structures. Violation of regional blood circulation of the lower limb of the animal was achieved by crossing the femoral artery. Limb immobilization was carried out for 4 weeks. The results were evaluated by the severity of changes in the morphological structure of the operated joints and radiographic examination. The high mitotic activity of cellular elements, revealed as a result of histological examination in all groups of animals, indicates the rapid formation of vascular collaterals in rats. It was noted that with non-rigid immobilization of the joint with a plaster cast, the cartilaginous tissue of the articular surface becomes thinner, while the subchondral structures change slightly. With severe immobilization of the joint in the external fixation apparatus, significant changes are revealed precisely in the structure of subchondral structures. In animals subjected to a two-week rehabilitation period after dismantling the external fixation apparatus, a further progression of the pathological process was noted. Thus, with severe immobilization of the limb in the experiment, it was possible to obtain a stable joint deformation corresponding to the 3rd clinical and radiological stage of osteoarthritis.

Background. Charcot arthropathy is a serious medical and social problem. Histological studies of foot joints components in Charcot arthropathy complicated by osteomyelitis are few. The purpose of this study was to assess structural changes in the articular cartilage and subchondral bone of the foot joints in Charcot arthropathy complicated by osteomyelitis. Materials and Methods. The bone-cartilage fragments of the ankle, subtalar and metatarsophalangeal joints with the surrounding soft tissues of 20 patients with Charcot arthropathy complicated by chronic osteomyelitis were examined. Part of the material was fixed in neutral formalin. All samples were subjected to standard histological processing. Paraffin sections were stained by Masson’s tricolor method, hematoxylin and eosin. Part of the material was embedded in epoxy resins. Then semi-thin sections were stained with methylene blue and basic fuchsin. Histological preparations were studied by digitizing images under the AxioScope A1 microscope (Carl Zeiss MicroImaging GmbH, Germany).The phase of chronic osteomyelitis inflammation was assessed semi-quantitatively using the histopathological scale by A. Tiemann et al. (2014). Results. In 80% of the patients, the inflammatory phase of chronic osteomyelitis was characterized as active and subacute. In all cases, the areas with full-layer of articular cartilage unweaving, up to the subchondral zone, with cartilaginous tissue fragments rejection into the joint cavity were revealed. Cytoarchitectonics was disrupted. The main part of chondrocytes was in a state of destruction. The articular surface was covered with pannus. There were no basophilic line and the zone of calcified cartilage. The hyaline cartilage was replaced by granulation and/or fibrous tissue. An inflammatory infiltrates was noted in the superficial and deep areas of the cartilage. The impairment of the structure and/or complete absence of the subchondral bone due to the high activity of osteoclasts in the subchondral zone were revealed. An excessive amount of osteoclasts at the border with the articular cartilage was noted, while the signs of reparative bone formation were poorly expressed. Edema and thickening of the vascular walls of the microvasculature were recorded. Conclusion. The microscopic examination of the foot joints in Charcot arthropathy complicated by osteomyelitis revealed structural impairment and/or complete absence of the subchondral bone due to the high activity of osteoclasts in the subchondral zone. Structural changes in the subchondral bone and synovial pannus led to irreversible destruction of articular cartilage and the penetration of infection. These should be taken into account in surgical planning.

Summary Objective Sprifermin (recombinant human fibroblast growth factor-18), a potential disease-modifying osteoarthritis (OA) drug, demonstrated dose-dependent effects on femorotibial cartilage thickness (by quantitative magnetic resonance imaging [MRI]) in the phase II FORWARD study. This post-hoc analysis evaluated the potential effects of sprifermin on several articular structures in the whole joint over 24 months using semi-quantitative MRI assessment. Design Patients aged 40–85 years with symptomatic radiographic knee OA, Kellgren–Lawrence grade 2 or 3, and medial minimum joint space width ≥2.5 mm in the target knee were randomized (1:1:1:1:1) to receive three double-blinded, once-weekly, intra-articular injections of sprifermin 30 μg or 100 μg or placebo every 6 (q6mo) or 12 months. 1.5- or 3 T MRIs were read using the Whole-Organ Magnetic Resonance Imaging Score (WORMS) system at baseline and 24 months. Change from baseline at 24 months on compartment and/or whole knee level was assessed for cartilage morphology, bone marrow lesions (BMLs), and osteophytes by delta-subregional and delta-sum (DSM) approaches. Menisci, Hoffa-synovitis, and effusion-synovitis were also evaluated for worsening. Results 549 patients were included. Dose-dependent treatment effects from baseline to 24 months were observed on cartilage morphology (sprifermin 100 μg q6mo vs placebo; mean DSM (95% confidence interval [CI]) −0.6 (−1.5, 0.2); less cartilage worsening) in the entire knee and BMLs sprifermin 100 μg q6mo vs placebo; mean DSM (95% CI) −0.2 (−0.5, 0.1) in the patellofemoral compartment. No effects over 24 months were observed on osteophytes, menisci, Hoffa-synovitis or effusion-synovitis. Conclusions Positive effects associated with sprifermin were observed for cartilage morphology changes, and BML improvement. There were no meaningful negative or positive effects associated with sprifermin in the other joint tissues examined.

Objectives Residual hip dysplasia is the most common underlying condition leading to secondary osteoarthritis (OA) of the hip. Subchondral bone alterations in OA secondary to hip dysplasia (HD-OA) are poorly investigated. The aim of the present study was to analyse the microarchitecture, bone remodelling and pathological alterations of subchondral bone in femoral heads from patients with HD-OA. Methods Subchondral bone specimens were extracted from both weight-bearing and non–weight-bearing regions of femoral heads from 20 patients with HD-OA and 20 patients with osteoporotic femoral neck fracture, during hip replacement surgery. Micro-CT and histological examination were performed to assess the microarchitecture and histopathological changes. Results The weight-bearing subchondral bone showed significantly more sclerotic microarchitecture and higher bone remodelling level in HD-OA as compared with osteoporosis. In the non–weight-bearing region, the two diseases shared similar microarchitectural characteristics, but higher bone remodelling level was detected in HD-OA. Distinct regional differences were observed in HD-OA, whereas the two regions exhibited similar characteristics in osteoporosis. In addition, HD-OA displayed more serious pathological alterations, including subchondral bone cyst, metaplastic cartilaginous tissue, bone marrow oedema and fibrous tissue, especially in the weight-bearing region. Conclusions Osteoarthritic deteriorations of subchondral bone induced by hip dysplasia spread throughout the whole joint, but exhibit region-dependent variations, with the weight-bearing region more seriously affected. Biomechanical stress might exert a pivotal impact on subchondral bone homeostasis in hip dysplasia. The translational potential of this article The histomorphometric findings in the project indicate an early intervention for the development of hip dysplasia in clinic.

The Cytokine-like 1 (Cytl1) is first identified in bone marrow cells and preferentially expressed in cartilaginous tissue, and showed chondrogenic effects in mesenchymal cells, not essential for cartilage or bone development as in Cytl1 knock-out mice but associated with cartilage inflammatory and destruction. Here, we show the involvement of Cytl1 in chondrogenesis. Using specified chondrogenic embryonic skeleton and adult cartilage, the Cytl1 gene expression was investigated with associated chondrogenic factors by quantitative RT-PCR. The effect of Cytl1 protein (rCytl1) on cultured chondrocytes to regulate expression of key factors and phenotypic markers was studied. The results revealed that Cytl1 was highly expressed in chondrogenic process in embryos and adult cartilage. The rCytl1 increased the expression of Sox9 and Col2α1 with stabilized Col1α1 in cultured chondrocytes (redifferentiation). The Cytl1 was expressed and involved at all stages of cartilage development. Furthermore, Cytl1 expression shared similar patterns as other chondrogenic factors, implying interactions with other factors in chondrogenic process. Cytl1 is involved in cartilage development and matrix homeostasis, which defines the dedifferentiation phenotype of chondrocytes, essential to forming of functional cartilage in both physiologic remodeling and pathologic regeneration.

The comparative qualitative morphological study of bone fracture healing using etidronate-based preparation free of lanthanoids and in conjugation with them. The fragments of bone tissue from the facture zone were studied. They were obtained in 45 mongrel white rats on day 7, day 14, and day 30 after osteotomy in the zone of femoral shaft middle third. The investigation was performed by a morphological method in the three groups: (1) control, a fracture without treatment; (2) after paraosseous infusion of etidronate with lanthanoid and calcium ions; (3) after paraosseous infusion of etidronate with calcium ions without lanthanoids. The areas of leukocyte-necrotic masses with blood clots, those of granulation, connective and cartilaginous tissue, coarse-fibrous trabecular bone and lamellar bone, as well as those of necrotic bone parts were determined in the histological sections of the fracture zone with a morphometric grid. The data obtained were processed statistically. The best results of bone tissue healing for fractures were obtained when paraosseous infusing etidronate in combination with lanthanoids, and the effect of this was revealed just in the early periods after trauma. In this case, the intensity of injury-induced inflammatory reaction lowered and the volume of leukocyte-necrotic masses and traumatic edema decreased. The process of reparative regeneration accelerated as well with formation of granulation tissue and after that connective tissue and coarse-fibrous trabecular bone. Besides, indirect osteogenesis with bone formation through cartilaginous tissue was less common. As a result, there was uncomplicated fracture healing in most observations with the presented significant amount of formed compact and spongy lamellar bone. The combination of the etidronate-based preparation with lanthanoids is the most optimal, and its use is effective at all stages of bone fracture healing.

Injectable hydrogels have attracted increasing attention because of convenient clinical operation, non-invasive surgical procedure and seamless filling of irregular defects. Here, injectable di-self-crosslinking HSMSSA hydrogel was formed via fast thiol/maleimide click chemistry reaction and thiol oxidation reaction as primary and secondary self-crosslinking network, respectively. Molecular weight and precursor concentration significantly affected physichemical properties and biological functions of hydrogels. Although single HSMSSA gel (0.1 M Da, 10 mg/mL) had moderate injectability, preferable mechanical properties and good proliferative ability of chondrocytes in vitro, and could greatly promote cartilaginous tissue formation in vivo, the lack of adhesion sites resulted in an untenable situation in maintaining effective connections among newborn cell clusters. However, the biomimetic injectable di-self-crosslinking blend hydrogel by combing injectable HSMSSA and bioactive Col I had improved resistance to degradation, chondrocytes adhesion and proliferation, especially for multiples ascending genes expression level associated with hyaline cartilage formation and polyproteoglycan secretion, which might be a potential clinical treatment strategy for constructing injectable cartilage repair filler by combining expanded autologous chondrocytes. Statement of Significance An injectable di-self-crosslinking Hyaluronan–Based hydrogel was formed via fast thiol/maleimide click chemistry reaction and thiol oxidation reaction as primary/secondary self-crosslinking network, respectively. Molecular weight and precursor concentration significantly affected physichemical properties and biological functions of the hydrogels. Although this HSMSSA gel (0.1 M Da, 10 mg/mL) had moderate injectability, preferable mechanical properties, and good proliferative ability of chondrocytes in vitro, and could greatly promote cartilaginous tissue formation in vivo, the lack of adhesion sites resulted in ineffective connections among newborn cell clusters. The biomimetic injectable di-self-crosslinking blend hydrogel improved chondrocyte adhesion and proliferation by combined injectable HSMSSA and bioactive Col I, especially for multiple ascending gene expression levels associated with hyaline cartilage formation and polyproteoglycan secretion.

Objective. To reveal the influence of functional mobility of connective-tissue structures on the development and course of intra-articular pathology of the knee joint. Materials and methods. Patients (n = 279) with intra-articular pathology of the knee joint after the arthroscopic treatment were examined. The state of functional mobility of connective-tissue structures was diagnosed using the express method for dysplastic state of connective tissues in conditions of ambulatory-polyclinic service. Statistical analysis was carried out with Pirson correlation criteria. Results. The paper is devoted to analysis of the influence of functional connective-tissue structure mobility on the development and course of intra-articular pathology of the knee joint. Subjects with low functional mobility of the connective-tissue structures were established to be more often subjected to chondromalacia of cartilaginous tissue of the femoral medial condyle and injury of the medial meniscus. Such pathology as slipping patella, Goff body hypertrophy and knee joint chondromatosis are typical for subjects with hypermobility of the connective-tissue structures. Conclusions. High degree of functional mobility of connective-tissue structures significantly contributes to preservation of cartilaginous cover of the femoral medial condyle, even with concomitant pathology of the knee joint available. For men with high functional mobility of connective-tissue structures, the most common diseases are knee joint chondromatosis and Goff body hypertrophy, and for women slipping patella. For women with low functional mobility of connective-tissue structures, the most typical is pathology of the medial meniscus.

Introduction Cutting the cost of manufacturing is important for extending the use of tissue-engineered therapeutic products. The present study aimed to develop a simple method for fabrication of cartilaginous tissues for regenerative therapy, utilizing the phenomenon where human articular chondrocytes grow thickness-wise and spontaneously form three-dimensionally thick tissues. Methods Normal human articular chondrocytes (NHACs) were cultured with varying concentrations of transforming growth factor beta 1 (TGF-β1) and/or fibroblast growth factor-2 (FGF-2) to optimize the culture condition for thickness-wise growth of chondrocytes. Next, the tissues grown in the optimal condition were subjected to re-differentiation culture in attached and detached states to assess differentiation capacity by evaluating secreted factors, histological analysis, and a gene expression assay. Results NHACs grew thickness-wise efficiently in the presence of 1 ng/mL TGF-β1 and 10 ng/mL FGF-2. After two weeks of culture, NHACs grew with 11-fold higher thickness and 16-fold higher cell number compared to cells which were neither treated with TGF-β1 nor with FGF-2. These thickness-wise-grown chondrocytes could be re-differentiated by a differentiation medium according to the increase in melanoma inhibitory activity (MIA) and positive safranin-O staining. Interestingly, the cartilaginous gene expression was considerably different between the attached and detached conditions even in the same culture medium, indicating the necessity of detachment and shrinkage to achieve further differentiation. Conclusions Spontaneous thickness-wise growth might provide a simple tissue-engineering method for manufacturing cartilaginous 3D tissues., Graphical abstract Image 1, Highlights • Chondrocytes grow thickness-wise efficiently in the presence of FGF-2 and TGF-β1 to medium. • Differentiation medium leads to MIA secretion and Safranin-O staining. • Detachment from culture surface promoted differentiation of chondrocytes.

The shortage of organ supply reduces the success rate of organ transplantation. Hence, tissue regeneration has been initiated with the intention of improving the available treatment modalities. Articular cartilage is a suitable tissue for this purpose due to its limited self-heal ability. This study aims to evaluate the cartilaginous properties of in vivo constructs formed using chondrocytes transfected with the combination of sry (sex determining region y)-box9 (SOX9) and telomerase reverse transcriptase (TERT) genes (SOX9/TERT-transfected chondrocytes) seeded on a three-dimensional (3D) poly(lactic-co-glycolic) acid (PLGA)-based scaffold. The rabbit’s articular chondrocytes (n=6) were transfected with SOX9 and TERT genes via lipofection. The non-transfected chondrocyte (NTC) was used as a control. A total of 1×105 cells were seeded on a PLGA and PLGA/fibrin hybrid scaffolds to form constructs. The resulted constructs were SOX9/TERT-PLGA/fibrin, NTC-PLGA/fibrin, SOX9/TERT-PLGA and, NTC-PLGA. All constructs were cultured for three weeks prior to subcutaneous implantation into the athymic mice for two and four weeks. The constructs’ structural and functional aspects were evaluated using macroscopic observation, compression-stress analysis, histology, quantitative sulphated glycosaminoglycan (sGAG) assay and cartilage-specific genes (ACAN, COL2A1, SOX9), TERT, and MMP13 expression analysis. The constructs demonstrated a cartilage-like appearance. The constructs’ rigidity corresponded to the homogenous cells and extracellular matrix distribution in the week-4 constructs. Correspondingly, the cartilaginous matrix components were visualised at the pericellular matrix region of the construct, supported by the increment of quantitative sGAG content. The SOX9/TERT-PLGA/fibrin exhibited better genes expression and cartilaginous phenotypes than the other construct groups. The SOX9/TERT-PLGA/fibrin construct facilitated cartilaginous tissue formation.

Humic acids are characterized by chelating properties due to which they are able to influence the mineral metabolism in animals. Rabbits have species-specific characteristics of mineral metabolism, which distinguishes them from most species of mammals. For the experiment, 16 rabbits of 45-day-old of Hyplus breed were divided into the experimental and control group of 8 rabbits in each. The animals were 15 days in the preparatory period. The rabbits of the experimental group received humic acid additive individually with water in the amount of 5 mg/kg, the rabbits of the control group received pure water in the same amount. The ability of humic acid additive to increase the content of calcium, ionized calcium, iron, to correct the content of copper and zinc, and also to increase the activity of alkaline phosphatase in the blood serum of rabbits of the experimental group has been proved. We observed the early effect of the humic acid additive on increasing the content of iron and calcium ionized and a later effect on the activity of alkaline phosphatase and an increase in the content of copper in the blood serum. The influence of humic acid additive on the distribution of calcium, phosphorus, manganese, copper and zinc in bone tissue is determined. According to a histological study, there was a significant increase in the number of layers of osteons and osteoblasts in the bone tissue of the femur, an increase in the number of osteons and osteoblasts in the bone tissue of the sternum and an increase in the number of columns of chondrocytes and an increase in the number of chondrocytes in the column in the cartilaginous tissue of the sternum. The results of a histological study of bone tissue, together with an increase in structural macroelements in it and redistribution of osteotropic trace elements in the blood and bone tissue suggest the possibility of using a humic acid additive to intensify the growth and mineralization of bone tissue, which can improve the state of bone tissue of rabbits in the postnatal period ontogenesis.

Background Although previous studies have suggested a stimulatory role of heparanase in physiological bone turnover, the potential therapeutic role of heparanase in bone healing has not been elucidated. The purpose of this study was to assess the effect of topical application of heparanase-1 on bone healing. Methods Two different dosages of recombinant mouse heparanase-1 and vehicle control were prepared and delivered via an osmotic pump to provide continuous topical infusion of the therapeutic reagent in a mouse bone defect model at the distal femoral metaphysis. The bone healing progress was evaluated by micro-computed tomography and histological examination at 7, 14, and 21 days after the bone defect was created. Results The peak of trabecular bone generation was achieved earlier than anticipated with the use of heparanase as measured by medullary bone volume fraction and trabecular number observed in micro-computed tomography, while the remodeling of trabecular bone to cortical bone was also achieved earlier than anticipated with the use of heparanase as measured by connectivity density. Histopathological observation revealed a higher frequency of the presence of cartilaginous tissue in the heparanase-treated groups. Both bone mineral density and cortical bone volume fraction showed the best healing outcome with low-dose heparanase, implying a biphasic effect of its mode of action. Conclusion These results indicated that with the appropriate dose of topical heparanase-1, the progress of bone healing could be accelerated in vivo.

The physis is a cartilaginous tissue in children's long bones that is responsible for bone elongation. Physeal injuries can heal with bony repair tissue known as a "bony bar," and this can cause growth deformities. Current treatments involve surgical resection of the bony bar and insertion of inert materials in hopes of preventing bony bar re-formation and preserving bone elongation. However, these materials frequently fail and the bony bar commonly returns. This study investigated alginate-chitosan hydrogels as interpositional materials to block bony bar formation in a rat model of physeal injury. Further, biomaterial properties such as substrate stiffness, permeability, and degradation rate were studied. Different ratio alginate:chitosan hydrogels with or without calcium cross-linking were tested for their inhibition of bony bar formation and restoration of the injured physis. Alginate:chitosan were mixed (a) 90:10 with calcium (90:10 + Ca); (b) 50:50 with calcium (50:50 + Ca); (c) 50:50 without calcium (50:50 - Ca); and (d) 50:50 made with irradiated alginate (IA) and without calcium. We found that repair tissue was determined primarily by the in vivo degradation rate of alginate-chitosan hydrogels. 90:10 + Ca had a slow degradation rate, prevented cellular infiltration, and produced the most bony bar tissue while having softer, more permeable material properties. IA had the fastest degradation, showed high cellular infiltration, and produced the most cartilage-like tissue while having stiffer, less permeable material properties. Our results suggest that the in vivo biomaterial degradation rate is a dynamic property that can be optimized to influence cell fate and tissue repair in physeal injuries.

Recently, many studies have focused on the repair and regeneration of damaged articular cartilage using tissue engineering. In tissue engineering therapy, cells are cultured in vitro to create a three-dimensional (3-D) tissue designed to replace the damaged cartilage. Although tissue engineering is a useful approach to regenerating cartilage, mechanical anisotropy has not been reconstructed from a cellular organization level. This study aims to create mechanically anisotropic cartilaginous tissue using dielectrophoretic cell patterning and gel-sheet lamination. Bovine chondrocytes were patterned in a hydrogel to form line-array cell clusters via negative dielectrophoresis (DEP). The results indicate that the embedded chondrocytes remained viable and reconstructed cartilaginous tissue along the patterned cell array. Moreover, the agarose gel, in which chondrocytes were patterned, demonstrated mechanical anisotropy. In summary, our DEP cell patterning and gel-sheet lamination techniques would be useful for reconstructing mechanically anisotropic cartilage tissues.

PURPOSE: The purpose of this study was to characterize the influence of SDF-1 on cell migration/adhesion and temporal gene expression of human cartilage mesenchymal progenitor cells (C-PCs); and to utilize SDF-1 conditioned mesenchymal progenitors to stimulate reintegration of human meniscus fibrocartilage breaks. MATERIALS AND METHODS: Characterization of SDF-1-induced cell migration was achieved using hydroxypropyl cellulose (HPC) scaffolds pretreated with SDF-1. Fluorescence microscopy and cell counting were used to visualize and quantify the extent of cell migration into scaffolds, respectively. Relative mRNA expression analysis was used to characterize the temporal effects of SDF-1 on C-PCs. Tissue reintegration experiments were conducted using cylindrical human meniscal tissue punches, which were then placed back together with an HPC scaffold embedded with C-PCs. Tensile testing was used to evaluate the extent of tissue reintegration stimulated by human mesenchymal progenitors. RESULTS: C-PCs migrate into scaffolds in response to SDF-1 with the same efficiency as mesenchymal progenitors from human marrow (BM-MSCs). SDF-1 treatment of C-PCs did not significantly alter the expression of early and late stage chondrogenic differentiation genes. Scaffolds containing SDF-1 preconditioned C-PCs successfully adhered to fibrocartilage breaks and migrated from the scaffold into the tissue. Tensile testing demonstrated that SDF-1 preconditioned C-PCs stimulate reintegration of fibrocartilage tears. CONCLUSION: C-PCs migrate in response to SDF-1. Exposure to SDF-1 does not significantly alter the unique mRNA profile of C-PCs that make them desirable for cartilaginous tissue repair applications. SDF-1 pretreated mesenchymal progenitors successfully disperse into injured tissues to help facilitate tissue reintegration.

Cartilage lacks basic repair mechanisms and thus surgical interventions are necessary to treat lesions. Minimally-invasive arthroscopic procedures require the development of injectable biomaterials to support chondrogenesis of implanted cells. However, most cartilage tissue engineering approaches rely on pre-culture of scaffolds in media containing growth factors (GFs) such as transforming growth factor (TGF)-β1, which are crucial for cartilage formation and homeostasis. GFs media-supplementation is incompatible with injectable approaches and has led to a knowledge gap about optimal dose of GFs and release profiles needed to achieve chondrogenesis. This study aims to determine the optimal loading and release kinetics of TGF-β1 bound to an engineered GAG hydrogel to promote optimal cartilaginous matrix production in absence of TGF-β1 media-supplementation. We show that heparin, a GAG known to bind a wide range of GFs, covalently conjugated to a hyaluronan hydrogel, leads to a sustained release of TGF-β1. Using this heparin-conjugated hyaluronan hydrogel, 0.25 to 50 ng TGF-β1 per scaffold was loaded and cell viability, proliferation and cartilaginous matrix deposition of the encapsulated chondroprogenitor cells were measured. Excellent chondrogenesis was found when 5 ng TGF-β1 per scaffold and higher were used. We also demonstrate the necessity of a sustained release of TGF-β1, as no matrix deposition is observed upon a burst release. In conclusion, our biomaterial loaded with an optimal initial dose of 5 ng/scaffold TGF-β1 is a promising injectable material for cartilage repair, with potentially increased safety due to the low, locally administered GF dose. Statement of Significance Cartilage cell-based products are dependent on exogenous growth factor supplementation in order for proper tissue maturation. However, for a one-step repair of defects without need for expensive tissue maturation, an injectable, growth factor loaded formulation is required. Here we show development of an injectable hyaluronan hydrogel, which achieves a sustained release of TGF-β1 due to covalent conjugation of heparin. These grafts matured into cartilaginous tissue in the absence of growth factor supplementation. Additionally, this system allowed us to screen TGF-β1 concentrations to determine the mimimum amount of growth factor required for chondrogenesis. This study represents a critical step towards development of a minimally-invasive, arthroscopic treatment for cartilage lesions.

Articular cartilage has limited capacity for regeneration and when damaged cannot be repaired with currently available metallic or synthetic implants. We aim to bioengineer a microfibre-reinforced hydrogel that can capture the zonal depth-dependent mechanical properties of native cartilage, and simultaneously support neo-cartilage formation. With this goal, a sophisticated bi-layered microfibre architecture, combining a densely distributed crossed fibre mat (superficial tangential zone, STZ) and a uniform box structure (middle and deep zone, MDZ), was successfully manufactured via melt electrospinning and combined with a gelatin–methacrylamide hydrogel. The inclusion of a thin STZ layer greatly increased the composite construct's peak modulus under both incongruent (3.2-fold) and congruent (2.1-fold) loading, as compared to hydrogels reinforced with only a uniform MDZ structure. Notably, the stress relaxation response of the bi-layered composite construct was comparable to the tested native cartilage tissue. Furthermore, similar production of sulphated glycosaminoglycans and collagen II was observed for the novel composite constructs cultured under mechanical conditioning w/o TGF-ß1 supplementation and in static conditions w/TGF-ß1 supplementation, which confirmed the capability of the novel composite construct to support neo-cartilage formation upon mechanical stimulation. To conclude, these results are an important step towards the design and manufacture of biomechanically competent implants for cartilage regeneration. Statement of Significance: Damage to articular cartilage results in severe pain and joint disfunction that cannot be treated with currently available implants. This study presents a sophisticated bioengineered bi-layered fibre reinforced cell-laden hydrogel that can approximate the functional mechanical properties of native cartilage. For the first time, the importance of incorporating a viable superficial tangential zone (STZ) – like structure to improve the load-bearing properties of bioengineered constructs, particularly when in-congruent surfaces are compressed, is demonstrated. The present work also provides new insights for the development of implants that are able to promote and guide new cartilaginous tissue formation upon physiologically relevant mechanical stimulation.