Cartilage Collagen Research Articles

Targeting FAP-positive chondrocytes in osteoarthritis: a novel lipid nanoparticle siRNA approach to mitigate cartilage degeneration

BackgroundOsteoarthritis (OA) is a common joint disease that leads to chronic pain and functional limitations. Recent research has revealed soluble fibroblast activation protein (FAP) secreted from OA synovium could degrade type II collagen (Col2) in cartilage to promote the progression of OA. This study aimed to reveal the role of FAP from chondrocytes in OA and develop a novel lipid nanoparticle (LNP)-FAP siRNA delivery system for OA treatment.MethodsThe expression of FAP in the cartilage of knee OA patients was investigated using [68 Ga]Ga-FAPI-04 PET in vivo and immunofluorescence, western blotting, and RT-qPCR in vitro. Cell senescence was determined by senescence-associated β-galactosidase (SA-β-Gal) assay after FAP overexpressing or knockdown in chondrocytes. An OA model with chondrocyte-specific FAP knockout mice was applied to investigate the role of FAP in chondrocyte senescence and OA development. The therapeutic effects of lipid nanoparticle (LNP) @FAP siRNA on cartilage degeneration were evaluated in the rat OA model.ResultsOur study found that higher [68 Ga]Ga-FAPI-04 uptake was detected in knee OA patients by PET/CT scan. FAP mRNA and protein levels were highly expressed in OA-damaged cartilage. Moreover, we found that overexpression of FAP promotes chondrocyte senescence, and the genetic knockout of FAP in chondrocytes alleviates OA. Knockdown FAP by siRNA could alleviate chondrocyte senescence and suppress the NF-κB pathway to reduce the senescence-associated secretory phenotype (SASP). In the rat model of OA, intraarticular injection of LNP@FAP siRNA can reduce senescent cells and ameliorate cartilage destruction.ConclusionFAP-positive chondrocytes play a significant role in the pathogenesis of OA. Targeting these cells selectively has the potential to mitigate the progression of the disease. Our study provides valuable insights into the intraarticular injection of LNP@FAP siRNA as a promising strategy for the treatment of OA.Graphical abstract

Cellular solids and prestressed affine networks as models of the elastic behavior of soft biological structures.

We reviewed two microstructural models, cellular solid models and prestressed affine network models, that have been used previously in studies of elastic behavior of soft biological materials. These models provide simple and mathematically transparent equations that can be used to interpret experimental data and to obtain quantitative predictions of the elastic properties of biological structures. In both models, volumetric density and elastic properties of the microstructure are key determinants of the macroscopic elastic properties. In the prestressed network model, geometrical rearrangement of the microstructure (kinematic stiffness) is also important. As examples of application of these models, we considered the shear behavior of the cytoskeleton of adherent cells, of the collagen network of articular cartilage, and of the lung parenchymal network since their ability to resist shear is important for their normal biological and physiological functions. All three networks carry a pre-existing stress (prestress). We predicted their shear moduli using the microstructural models and compared those predictions with existing experimental data. Prestressed network models of the cytoskeleton and of the lung parenchyma provided a better correspondence to experimental data than cellular solid models. Both cellular solid and prestressed network models of the cartilage collagen network provided reasonable agreements with experimental values. These findings suggested that the kinematic stiffness and material stiffness of microstructural elements were both important determinants of the shear modulus of the cytoskeleton and of the lung parenchyma, whereas elasticity of collagen fibrils had a predominant role in the cartilage shear behavior.

Hesperetin Attenuates T-2 Toxin-Induced Chondrocyte Injury by Inhibiting the p38 MAPK Signaling Pathway.

Hesperetin, a flavonoid derived from citrus fruits, exhibits potent antioxidant and anti-inflammatory activities and has been implicated in cartilage protection. However, its effectiveness against T-2 toxin-induced knee cartilage damage remains unclear. In this study, high-throughput sequencing analysis was employed to identify the key signaling pathways involved in T-2 toxin-induced articular cartilage damage in rats. Animal models were divided into the following groups: control, low-dose T-2 toxin, high-dose T-2 toxin, T-2 toxin + hesperetin, hesperetin, and vehicle. Pathological staining and immunohistochemistry were used to assess pathological changes, as well as the expression levels of the cartilage matrix-related proteins MMP13 and collagen II, along with the activation of the p38 MAPK signaling pathway. Additionally, primary rat chondrocytes were cultured to establish an in vitro model for investigating the underlying mechanism. High-throughput sequencing analysis revealed the involvement of the MAPK signaling pathway in T-2 toxin-induced articular cartilage damage in rats. Hesperetin intervention in T-2 toxin-exposed rats attenuated pathological cartilage damage. Immunohistochemistry results demonstrated a significant reduction in collagen II protein expression in the high-dose T-2 toxin group (p < 0.01), accompanied by a significant increase in MMP13 protein expression (p < 0.01). In both the articular cartilage and the epiphyseal plate, the T-2 toxin + hesperetin group exhibited significantly higher collagen II protein expression than the high-dose T-2 toxin group (p < 0.05), along with significantly lower MMP13 protein expression (p < 0.05). Hesperetin inhibited the over-activation of the p38/MEF2C signaling axis induced by T-2 toxin in primary rat chondrocytes. Compared to the T-2 toxin group, the T-2 toxin + hesperetin group showed significantly reduced phosphorylation levels of p38 and protein expression levels of MEF2C (p < 0.001 or p < 0.05). Moreover, the T-2 toxin + hesperetin group exhibited a significant decrease in MMP13 protein expression (p < 0.05) and a significant increase in collagen II protein expression (p < 0.01) compared to the T-2 toxin group. T-2 toxin activates the p38 MAPK signaling pathway, causing knee cartilage damage in rats. Treatment with hesperetin inhibits the p38/MEF2C signaling axis, regulates collagen II and MMP13 protein expression, and reduces cartilage injury significantly.

Protective Effects of Vitamin D on Proteoglycans of Human Articular Chondrocytes through TGF-β1 Signaling.

The extracellular matrix of cartilage primarily constitutes of collagen and aggrecan. Cartilage degradation starts with aggrecan loss in osteoarthritis (OA). Vitamin D (VD) plays an essential role in several inflammation-related diseases and can protect the collagen in cartilage during OA. The present study focused on the role of VD in aggrecan turnover of human articular chondrocytes treated with tumor necrosis factor α (TNF-α) and the possible mechanism. Treatment with different doses of VD and different periods of intervention with TNF-α and TGF-β1 receptor (TGFβR1) inhibitor SB525334 were investigated. The viability of human chondrocytes and extracellular secretion of TGF-β1 were measured. The expression of intracellular TGFβR1 and VD receptor was examined. Transcriptional and translational levels of aggrecan and the related metabolic factors were analyzed. The results showed that TNF-α markedly reduced the viability, TGFβR1 expressions and aggrecan levels of human chondrocytes, and increased disintegrin and metalloproteinase with thrombospondin motifs. The alterations were partially inhibited by VD treatment. Furthermore, the effects of VD were blocked by the TGFβR1 inhibitor SB525334 in TNF-α-treated cells. VD may prevent proteoglycan loss due to TNF-α via TGF-β1 signaling in human chondrocytes.

Changes of bone and articular cartilage in broilers with femoral head necrosis

Femoral head necrosis (FHN) in broilers is a common leg disorder in intensive poultry farming, giving rise to poor animal health and welfare. Abnormal mechanical stress in the hip joint is a risk factor for FHN, and articular cartilage is attracting increasing attention as a cushion and lubrication structure for the joint. In the present study, broilers aged 3 to 4 wk with FHN were divided into femoral head separation (FHS) and femoral head separation with growth plate lacerations (FHSL) groups, with normal broilers as control. The features of the hip joint, bone, and cartilage were assessed in FHN progression using devices including computed tomography (CT), atomic force microscope (AFM), and transmission electron microscopy (TEM). Broilers with FHN demonstrated decreased bone mechanical properties, narrow joint space, and thickened femoral head stellate structures. Notably, abnormal cartilage morphology was observed in FHN-affected broilers, characterized by increased cartilage thickness and rough cartilage surfaces. In addition, as FHN developed, cartilage surface friction and friction coefficient dramatically increased, while cartilage modulus and stiffness decreased. The ultramicro-damage occurred in chondrocytes and the extracellular matrix (ECM) of cartilage. Cell disintegration, abnormal mitochondrial accumulation, and oxidative stress damage were observed in chondrocytes. A notable decline in cartilage collagen content was observed in ECM during the initial stages of FHN, accompanied by a pronounced reduction in collagen fiber diameter and proteoglycan content as FHN progressed. Furthermore, the noticeable loosening of the collagen fiber structure and the appearance of type I collagen were noted in cartilage. In conclusion, there was a progressive decrease in bone quality and multifaceted damage of cartilage in the femoral head, which was closely linked to the severity of FHN in broilers.

ER procollagen storage defect without coupled unfolded protein response drives precocious arthritis.

Collagenopathies are a group of clinically diverse disorders caused by defects in collagen folding and secretion. For example, mutations in the gene encoding collagen type-II, the primary collagen in cartilage, can lead to diverse chondrodysplasias. One example is the Gly1170Ser substitution in procollagen-II, which causes precocious osteoarthritis. Here, we biochemically and mechanistically characterize an induced pluripotent stem cell-based cartilage model of this disease, including both hetero- and homozygous genotypes. We show that Gly1170Ser procollagen-II is notably slow to fold and secrete. Instead, procollagen-II accumulates intracellularly, consistent with an endoplasmic reticulum (ER) storage disorder. Likely owing to the unique features of the collagen triple helix, this accumulation is not recognized by the unfolded protein response. Gly1170Ser procollagen-II interacts to a greater extent than wild-type with specific ER proteostasis network components, consistent with its slow folding. These findings provide mechanistic elucidation into the etiology of this disease. Moreover, the easily expandable cartilage model will enable rapid testing of therapeutic strategies to restore proteostasis in the collagenopathies.

Time-Dependent Effect of Eggshell Membrane on Monosodium-Iodoacetate-Induced Osteoarthritis: Early-Stage Inflammation Control and Late-Stage Cartilage Protection.

Osteoarthritis (OA) is a chronic degenerative joint disease that causes chronic pain, swelling, stiffness, disability, and significantly reduces the quality of life. Typically, OA is treated using painkillers and non-steroidal anti-inflammatory drugs (NSAIDs). While current pharmacologic treatments are common, their potential side effects have prompted exploration into functional dietary supplements. Recently, eggshell membrane (ESM) has emerged as a potential functional ingredient for joint and connective tissue disorders due to its clinical efficacy in relieving joint pain and stiffness. Despite promising clinical evidence, the effects of ESM on OA progression and its mechanism of action remain poorly understood. This study evaluated the efficacy of Ovomet®, a powdered natural ESM, against joint pain and disease progression in a monosodium iodoacetate (MIA)-induced rodent model of OA in mice and rats. The results demonstrate that ESM significantly alleviates joint pain and attenuates articular cartilage destruction in both mice and rats that received oral supplementation for 5 days prior to OA induction and for 28 days thereafter. Interestingly, ESM significantly inhibited mRNA expression levels of pro-inflammatory cytokines including tumor necrosis factor alpha (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6), as well as inflammatory mediators, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase in the knee joint cartilage at the early stage of OA, within 7 days after OA induction. However, this effect was not observed in the late stage at 28 days after OA induction. ESM further attenuates the induction of protein expression for cartilage-degrading enzymes like matrix metalloproteinase (MMPs) 3 and 13, and a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS-5), in the late-stage. In addition, MIA-induced reduction of the protein expression levels of cartilage components, cartilage oligomeric matrix protein (COMP), aggrecan (ACAN) and collagen type II α-1 chain (COL2α1), and cartilage extracellular matrix (ECM) synthesis promoting transcriptional factor SRY-Box 9 (SOX-9) were increased via ESM treatment in the cartilage tissue. Our findings suggest that Ovomet®, a natural ESM powder, is a promising dietary functional ingredient that can alleviate pain, inflammatory response, and cartilage degradation associated with the progression of OA.

Sensitivity of cartilage mechanical behaviour to spatial variations in material properties

Articular cartilage tissue exhibits a spatial dependence in material properties that govern mechanical behaviour. A mathematical model of cartilage tissue under one dimensional confined compression testing is developed for normal tissue that takes account of these variations in material properties. Modifications to the model representative of a selection of mechanisms driving osteoarthritic cartilage are proposed, allowing application of the model to both physiological and pathophysiological, osteoarthritic tissue. Incorporating spatial variations into the model requires the specification of more parameters than are required in the absence of these variations. A global sensitivity analysis of these parameters is implemented to identify the dominant mechanisms of mechanical response, in normal and osteoarthritic cartilage tissue, to both static and dynamic loading. The most sensitive parameters differ between dynamic and static mechanics of the cartilage, and also differ between physiological and osteoarthritic pathophysiological cartilage. As a consequence changes in cartilage mechanics in response to alterations in cartilage structure are predicted to be contingent on the nature of loading and the health, or otherwise, of the cartilage. In particular the mechanical response of cartilage, especially deformation, is predicted to be much more sensitive to cartilage stiffness in the superficial zone given the onset of osteoarthritic changes to material properties, such as superficial zone increases in permeability and reductions in fixed charge. In turn this indicates that any degenerative changes in the stiffness associated with the superficial cartilage collagen mesh are amplified if other elements of osteoarthritic disease are present, which provides a suggested mechanism-based explanation for observations that the range of mechanical parameters representative of normal and osteoarthritic tissue can overlap substantially.

Cartilage Collagen Neoepitope C2C Expression in the Articular Cartilage and Its Relation to Joint Tissue Damage in Patients with Knee Osteoarthritis.

Pathological cleavage of type II collagen (Col2) and generation of Col2 neoepitopes can serve as useful molecular markers of the progression of osteoarthritis (OA). One of such potential biomarkers is type II collagen neoepitope C2C. The aim of this study was to correlate the degree of articular cartilage damage in OA patients with C2C expression in histological samples of tissues removed during total knee replacement. Cartilage samples were obtained from 27 patients ranging in age from 55 to 66 years. In each patient, medial and lateral tibia plateau samples were analyzed according to the OARSI histopathology grading system. The C2C expression was evaluated on histological slides by semi-quantitative analysis using ImageJ Fiji 2.14.0 software. Spearman's rank correlation analysis revealed a positive weak correlation (rho = 0.289, p = 0.0356) between the histological grade of tissue damage and the percentage of C2C staining. In addition, a highly significant positive correlation (rho = 0.388, p = 0.0041) was discovered between the osteoarthritis score (combining the histological grade of damage with the OA macroscopic stage) and the percentage of C2C staining in the samples. The C2C expression was detected in all the regions of the articular cartilage (i.e., the superficial zone, mid zone, deep zone and tidemark area, and the zone of calcified cartilage). Our findings imply that local expression of C2C correlates with the articular cartilage damage in OA-affected knees. This confirms that C2C can be used as a prospective marker for assessing pathological changes in the OA course and OA clinical trials.

Ammodaucus Leucotrichus Seed Extract as a Potential Therapy in Animal Models of Rheumatoid Arthritis Induced by Complete Freund Adjuvant and Chicken Cartilage Collagen.

This study assessed the efficacy of an Ammodaucus leucotrichus seed extract to treat rheumatoid arthritis in rat models of this disease. Rheumatoid arthritis was induced in rats using two methods: immunization with 100 µL of Complete Freund Adjuvant (CFA) and immunization with 100 µL of a 3mg/ml solution of type II collagen (CII) from chicken cartilage. The therapeutic potential of the extract was assessed at different doses (150, 300, and 600mg/kg/day for 21 days in the CII-induced arthritis model and for 14 days in the CFA-induced arthritis model) and compared with methotrexate (MTX; 0.2mg/kg for the same periods), a commonly used drug for rheumatoid arthritis treatment in humans. In both models (CII-induced arthritis and CFA-induced arthritis), walking distance, step length, intra-step distance and footprint area were improved following treatment with the A. leucotrichus seed extract (all concentrations) and MTX compared with untreated animals. Both treatments increased the serum concentration of glutathione and reduced that of complement C3, malondialdehyde and myeloperoxidase. Radiographic data and histological analysis indicated that cartilage destruction was reduced already with the lowest dose of the extract (100mg/kg/dose) in both models. These results show the substantial antiarthritic potential of the A. leucotrichus seed extract, even at the lowest dose, suggesting that it may be a promising alternative therapy for rheumatoid arthritis and joint inflammation. They also emphasize its efficacy at various doses, providing impetus for more research on this extract as a potential therapeutic agent for arthritis.

Functional and Molecular Analysis of Human Osteoarthritic Chondrocytes Treated with Bone Marrow-Derived MSC-EVs.

Osteoarthritis (OA) is a degenerative joint disease, causing impaired mobility. There are currently no effective therapies other than palliative treatment. Mesenchymal stromal cells (MSCs) and their secreted extracellular vesicles (MSC-EVs) have shown promise in attenuating OA progression, promoting chondral regeneration, and modulating joint inflammation. However, the precise molecular mechanism of action driving their beneficial effects has not been fully elucidated. In this study, we analyzed MSC-EV-treated human OA chondrocytes (OACs) to assess viability, proliferation, migration, cytokine and catabolic protein expression, and microRNA and mRNA profiles. We observed that MSC-EV-treated OACs displayed increased metabolic activity, proliferation, and migration compared to the controls. They produced decreased proinflammatory (Il-8 and IFN-γ) and increased anti-inflammatory (IL-13) cytokines, and lower levels of MMP13 protein coupled with reduced expression of MMP13 mRNA, as well as negative microRNA regulators of chondrogenesis (miR-145-5p and miR-21-5p). In 3D models, MSC-EV-treated OACs exhibited enhanced chondrogenesis-promoting features (elevated sGAG, ACAN, and aggrecan). MSC-EV treatment also reversed the pathological impact of IL-1β on chondrogenic gene expression and extracellular matrix component (ECM) production. Finally, MSC-EV-treated OACs demonstrated the enhanced expression of genes associated with cartilage function, collagen biosynthesis, and ECM organization and exhibited a signature of 24 differentially expressed microRNAs, associated with chondrogenesis-associated pathways and ECM interactions. In conclusion, our data provide new insights on the potential mechanism of action of MSC-EVs as a treatment option for early-stage OA, including transcriptomic analysis of MSC-EV-treated OA, which may pave the way for more targeted novel therapeutics.

HIF-1α in cartilage homeostasis, apoptosis, and glycolysis in mice with steroid-induced osteonecrosis of the femoral head.

With the prevalence of coronavirus disease 2019, the administration of glucocorticoids (GCs) has become more widespread. Treatment with high-dose GCs leads to a variety of problems, of which steroid-induced osteonecrosis of the femoral head (SONFH) is the most concerning. Since hypoxia-inducible factor 1α (HIF-1α) is a key factor in cartilage development and homeostasis, it may play an important role in the development of SONFH. In this study, SONFH models were established using methylprednisolone (MPS) in mouse and its proliferating chondrocytes to investigate the role of HIF-1α in cartilage differentiation, extracellular matrix (ECM) homeostasis, apoptosis and glycolysis in SONFH mice. The results showed that MPS successfully induced SONFH in vivo and vitro, and MPS-treated cartilage and chondrocytes demonstrated disturbed ECM homeostasis, significantly increased chondrocyte apoptosis rate and glycolysis level. However, compared with normal mice, not only the expression of genes related to collagens and glycolysis, but also chondrocyte apoptosis did not demonstrate significant differences in mice co-treated with MPS and HIF-1α inhibitor. And the effects observed in HIF-1α activator-treated chondrocytes were similar to those induced by MPS. And HIF-1α degraded collagens in cartilage by upregulating its downstream target genes matrix metalloproteinases. The results of activator/inhibitor of endoplasmic reticulum stress (ERS) pathway revealed that the high apoptosis rate induced by MPS was related to the ERS pathway, which was also affected by HIF-1α. Furthermore, HIF-1α affected glucose metabolism in cartilage by increasing the expression of glycolysis-related genes. In conclusion, HIF-1α plays a vital role in the pathogenesis of SONFH by regulating ECM homeostasis, chondrocyte apoptosis, and glycolysis.

Preoperative Synovial Tissue and Synovial Fluid Biomarkers as Predictors for Outcomes After Knee Arthroscopy and ACL Reconstruction: A Narrative Review.

Biomarkers collected in synovial tissue and fluid have been identified as potential predictors of outcomes after arthroscopy. To provide a narrative review of the current literature that assesses the associations between preoperative biomarkers in the synovial fluid or synovial tissue and patient outcomes after knee arthroscopy. Narrative review. We searched the PubMed database with keywords, "biomarkers AND arthroscopy," "biomarkers AND anterior cruciate ligament reconstruction," and "biomarkers AND meniscectomy." To be included, studies must have collected synovial fluid or synovial tissue from patients before or during arthroscopic knee surgery and analyzed the relationship of biomarkers to postoperative patient outcomes. Biomarkers were classified into 4 main categories: metabolism of aggrecan in cartilage, metabolism of collagen in cartilage (type II collagen), noncollagenous proteins in the knee, and other. When biomarker levels and outcomes were expressed with continuous variables, we abstracted the Pearson or Spearman correlation coefficients as the effect measure. If the biomarker values were continuous and the outcomes binary, we abstracted the mean or median biomarker values in those with favorable versus unfavorable outcomes. We calculated effect sizes as the difference between means of both groups divided by the standard deviation from the mean in the group with better outcomes. Eight studies were included in the review. Each study reported different patient outcomes. Biomarkers associated with metabolism of aggrecan, type II collagen metabolism, and noncollagenous proteins as well as inflammatory biomarkers had statistically significant associations with a range of patient outcomes after knee arthroscopy. Difference across studies in sample size and outcome measures precluded choosing a single biomarker that best predicted patient outcomes. The findings suggest that biomarkers associated with metabolism of aggrecan, type II collagen metabolism, noncollagenous proteins, as well as inflammatory biomarkers may help surgeons and their patients anticipate surgical outcomes.

Effects of Leukocyte-rich platelet-rich plasma and Leukocyte-poor platelet-rich plasma on cartilage in a rabbit osteoarthritis model.

Osteoarthritis is a prevalent chronic disease. One of its primary pathological processes involves the degeneration of articular cartilage. Platelet-rich plasma (PRP) contains cytokines and growth factors that can stimulate the repair and regeneration of articular cartilage tissues. PRP may also slow the progression of osteoarthritis. The purpose of this experiment is to compare the efficacy of Leukocyte poor (LP) - PRP and Leukocyte rich (LR) - PRP in treating rabbit osteoarthritis and to investigate their mechanisms of action. Analyzing the impact of leukocytes on PRP therapeutic effectiveness will provide a valuable clinical reference for the choice of which PRP is better for the treatment of osteoarthritis. A rabbit osteoarthritis model was established by injecting papain into the knee joint cavity, and LP-PRP and LR-PRP were prepared through different centrifugation methods for injection into the knee joint cavity. Eight weeks after injection, rabbit knee cartilage specimens were observed for gross changes, HE staining, senna O-solid green staining, and immunohistochemistry of type II collagen and were quantitatively compared using Pelletier's score, Mankin's pathology score, and ImageJ image processing software. Injection of papain into the knee joint cavity successfully established a rabbit model of osteoarthritis. All three evaluation indexes differed significantly from those of the blank group (P<0.05). LP-PRP and LR-PRP exhibited therapeutic effects when compared with the model group. The two PRP groups had similar gross tissue appearance and pathology (P>0.05). The LR-PRP group had higher collagen type-II expression (P < 0.05) than the LP-PRP group. Both LP-PRP and LR-PRP proved therapeutic for the rabbit papain osteoarthritis model. The difference in leukocyte content between the two groups did not yield different cartilage morphology or other factors by 8 weeks posttreatment. LR-PRP displayed the ability to release more factors relevant to the metabolism of type II collagen than LP-PRP, enabling the preservation of into cartilage collagen content of type II collagen and delaying osteoarthritis progression.

MESENCHYMAL STEM CELLS MAINTAIN ARTICULAR STABILIZATION AND PROMOTE ENDOGENOUS CARTILAGE REPAIR AFTER HIGH TIBIAL OSTEOTOMY: A SECOND-LOOK ARTHROSCOPY STUDY

Varus malalignment increases the susceptibility of cartilage to mechanical overloading, which stimulates catabolic metabolism to break down the extracellular matrix and lead to osteoarthritis (OA). The altered mechanical axis from the hip, knee to ankle leads to knee joint pain and ensuing cartilage wear and deterioration, which impact millions of the aged population. Stabilization of the remaining damaged cartilage, and prevention of further deterioration, could provide immense clinical utility and prolong joint function. Our previous work showed that high tibial osteotomy (HTO) could shift the mechanical stress from an imbalanced status to a neutral alignment. However, the underlying mechanisms of endogenous cartilage stabilization after HTO remain unclear. We hypothesize that cartilage-resident mesenchymal stem cells (MSCs) dampen damaged cartilage injury and promote endogenous repair in a varus malaligned knee. The goal of this study is to further examine whether HTO-mediated off-loading would affect human cartilage-resident MSCs' anabolic and catabolic metabolism. This study was approved by IACUC at Xi'an Jiaotong University. Patients with medial compartment OA (52.75±6.85 yrs, left knee 18, right knee 20) underwent open-wedge HTO by the same surgeons at one single academic sports medicine center. Clinical data was documented by the Epic HIS between the dates of April 2019 and April 2022 and radiographic images were collected with a minimum of 12 months of follow-up. Medial compartment OA with/without medial meniscus injury patients with unilateral Kellgren /Lawrence grade 3–4 was confirmed by X-ray. All incisions of the lower extremity healed well after the HTO operation without incision infection. Joint space width (JSW) was measured by uploading to ImageJ software. The Knee injury and Osteoarthritis Outcome Score (KOOS) toolkit was applied to assess the pain level. Outerbridge scores were obtained from a second-look arthroscopic examination. RNA was extracted to quantify catabolic targets and pro-inflammatory genes (QiaGen). Student's t test for two group comparisons and ANOVA analysis for differences between more than 2 groups were utilized. To understand the role of mechanical loading-induced cartilage repair, we measured the serial changes of joint space width (JSW) after HTO for assessing the state of the cartilage stabilization. Our data showed that HTO increased the JSW, decreased the VAS score and improved the KOOS score significantly. We further scored cartilage lesion severity using the Outerbridge classification under a second-look arthroscopic examination while removing the HTO plate. It showed the cartilage lesion area decreased significantly, the full thickness of cartilage increased and mechanical strength was better compared to the pre-HTO baseline. HTO dampened medial tibiofemoral cartilage degeneration and accelerate cartilage repair from Outerbridge grade 2 to 3 to Outerbridge 0 to 1 compared to untreated varus OA. It suggested that physical loading was involved in HTO-induced cartilage regeneration. Given that HTO surgery increases joint space width and creates a physical loading environment, we hypothesize that HTO could increase cartilage composition and collagen accumulation. Consistent with our observation, a group of cartilage-resident MSCs was identified. Our data further showed decreased expression of RUNX2, COL10 and increased SOX9 in MSCs at the RNA level, indicating that catabolic activities were halted during mechanical off-loading. To understand the role of cartilage-resident MSCs in cartilage repair in a biophysical environment, we investigated the differentiation potential of MSCs under 3-dimensional mechanical loading conditions. The physical loading inhibited catabolic markers (IL-1 and IL-6) and increased anabolic markers (SOX9, COL2).Knee-preserved HTO intervention alleviates varus malalignment-related knee joint pain, improves daily and recreation function, and repairs degenerated cartilage of medial compartment OA. The off-loading effect of HTO may allow the mechanoregulation of cartilage repair through the differentiation of endogenous cartilage-derived MSCs.

Cationic tantalum oxide nanoparticle contrast agent for micro computed tomography reveals articular cartilage proteoglycan distribution and collagen architecture alterations

ObjectiveCationic tantalum oxide nanoparticles (Ta2O5-cNPs), as a newly introduced contrast agent for computed tomography (CT) of cartilage, offer quantitative evaluation of proteoglycan content and biomechanical properties. However, knowledge on the depth-wise impact of cartilage constituents on NP diffusion, particularly the influence of the collagen network, is lacking. In this study, we aim to establish the depth-dependent relationship between Ta2O5-cNP diffusion and cartilage constituents (proteoglycan content, collagen content and network architecture). MethodsOsteochondral samples (n = 30) were harvested from healthy equine stifle joints (N = 15) and the diffusion of 2.55 nm diameter cationic Ta2O5-cNPs into the cartilage was followed with µCT imaging for up to 96 hours. The diffusion-related parameters, Ta2O5-cNP maximum partition (Pmax) and diffusion time constant, were compared against biomechanical and depth-wise structural properties. Biomechanics were assessed using a stress-relaxation and sinusoidal loading protocols, whereas proteoglycan content, collagen content and collagen network architecture were determined using digital densitometry, Fourier-transform infrared spectroscopy and polarized light microscopy, respectively. ResultsThe Pmax correlates with the depth-wise distribution of proteoglycans (bulk Spearman’s ρ = 0.87, p < 0.001). More open collagen network architecture at the superficial zone enhances intake of Ta2O5-cNPs, but collagen content overall decreases the intake. The Pmax values correlate with the equilibrium modulus (ρ = 0.80, p < 0.001) of articular cartilage. ConclusionThis study establishes the feasibility of Ta2O5-cNPs for the precise and comprehensive identification of biomechanical and structural changes in articular cartilage via contrast-enhanced µCT.

The development of a nucleus pulposus-derived cartilage analog scaffold for chondral repair and regeneration.

Focal chondral defects (FCDs) significantly impede quality of life for patients and impose severe economic costs on society. One of the most promising treatment options-autologous matrix-induced chondrogenesis (AMIC)-could benefit from a scaffold that contains both of the primary cartilage matrix components-sulfated glycosaminoglycans (sGAGs) and collagen type II. Here, 17 different protocols were evaluated to determine the most optimum strategy for decellularizing (decelling) the bovine nucleus pulposus (bNP) to yield a natural biomaterial with a cartilaginous constituency. The resulting scaffold was then characterized with respect to its biochemistry, biomechanics and cytocompatibility. Results indicated that the optimal decell protocol involved pre-crosslinking the tissue prior to undergoing decell with trypsin and Triton X-100. The residual DNA content of the scaffold was found to be 32.64 ± 9.26 ng/mg dry wt. of tissue with sGAG and hydroxyproline (HYP) contents of 72.53 ± 16.43. and 78.38 ± 8.46 μg/mg dry wt. respectively. The dynamic viscoelastic properties were found to be preserved (complex modulus: 17.92-16.62 kPa across a range of frequencies) while the equilibrium properties were found to have significantly decreased (aggregate modulus: 11.51 ± 9.19 kPa) compared to the non-decelled fresh bNP tissue. Furthermore, the construct was also found to be cytocompatible with bone marrow stem cells (BMSCs). While it was not permissive of cellular infiltration, the BMSCs were still found to have lined the laser drilled channels in the scaffold. Taken together, the biomaterial developed herein could be a valuable addition to the AMIC family of scaffolds or serve as an off-the-shelf standalone option for cartilage repair.

Collagen-Based Hydrogels for Cartilage Regeneration.

Cartilage regeneration remains difficult due to a lack of blood vessels. Degradation of the extracellular matrix (ECM) causes cartilage defects, and the ECM provides the natural environment and nutrition for cartilage regeneration. Until now, collagen hydrogels are considered to be excellent material for cartilage regeneration due to the similar structure to ECM and good biocompatibility. However, collagen hydrogels also have several drawbacks, such as low mechanical strength, limited ability to induce stem cell differentiation, and rapid degradation. Thus, there is a demanding need to optimize collagen hydrogels for cartilage regeneration. In this review, we will first briefly introduce the structure of articular cartilage and cartilage defect classification and collagen, then provide an overview of the progress made in research on collagen hydrogels with chondrocytes or stem cells, comprehensively expound the research progress and clinical applications of collagen-based hydrogels that integrate inorganic or organic materials, and finally present challenges for further clinical translation.

Comparative assessment of the morphology and antigenicity of human osteochondral units using formalin and coagulant fixatives

Fixation is crucial for preserving tissue integrity during processing, and the most commonly used cross-linking fixative in immunohistochemistry is neutral buffered formalin, which requires antigen retrieval as a crucial step. The successful use of newer coagulant fixatives like methacarn and EMA to preserve isotopes and eliminate the need for antigen retrieval has been reported recently, but their role in decalcified osteochondral unit samples remains unknown. Limited information on the use of coagulant fixatives as formalin substitutes makes it important to comparatively evaluate their effects on osteochondral samples and the impact of antigen retrieval on different days. Osteochondral units from a patient with osteoarthritis who underwent total knee-replacement surgery were fixed with three studied fixatives (Formalin, Methacarn, EMA) and divided into four portions, for different time periods (Day 1, 3, 7, 10). Sections were decalcified, stained with Safranin O, Alcian Blue, Toluidine Blue, PicroSirius Red, Hematoxylin and Eosin, and immunohistochemical analysis of Collagen type II and type X with and without antigen retrieval was conducted. Formalin showed better hematoxylin uptake than coagulant-based fixatives, while all fixatives preserved tissue morphology without necrosis or cellular degeneration with comparable staining quality. Methacarn and EMA-fixed tissues showed higher uptake of collagen type II compared to formalin-fixed tissues, with collagen type II uptake occurring only in the cartilage region and collagen type X uptake occurring only in the bone region. The study highlights the effectiveness of methacarn and EMA as efficient alternatives to formalin, preserving tissue morphology and antigen specificity.

An injectable and 3D printable pro-chondrogenic hyaluronic acid and collagen type II composite hydrogel for the repair of articular cartilage defects

Current treatments for repairing articular cartilage defects are limited. However, pro-chondrogenic hydrogels formulated using articular cartilage matrix components (such as hyaluronic acid (HA) and collagen type II (Col II)), offer a potential solution if they could be injected into the defect via minimally invasive arthroscopic procedures, or used as bioinks to 3D print patient-specific customised regenerative scaffolds—potentially combined with cells. However, HA and Col II are difficult to incorporate into injectable/3D printable hydrogels due to poor physicochemical properties. This study aimed to overcome this by developing an articular cartilage matrix-inspired pro-chondrogenic hydrogel with improved physicochemical properties for both injectable and 3D printing (3DP) applications. To achieve this, HA was methacrylated to improve mechanical properties and mixed in a 1:1 ratio with Col I, a Col I/Col II blend or Col II. Col I possesses superior mechanical properties to Col II and so was hypothesised to enhance hydrogel mechanical properties. Rheological analysis showed that the pre-gels had viscoelastic and shear thinning properties. Subsequent physicochemical analysis of the crosslinked hydrogels showed that Col II inclusion resulted in a more swollen and softer polymer network, without affecting degradation time. While all hydrogels exhibited exemplary injectability, only the Col I-containing hydrogels had sufficient mechanical stability for 3DP applications. To facilitate 3DP of multi-layered scaffolds using methacrylated HA (MeHA)-Col I and MeHA-Col I/Col II, additional mechanical support in the form of a gelatin slurry support bath freeform reversible embedding of suspended hydrogels was utilised. Biological analysis revealed that Col II inclusion enhanced hydrogel-embedded MSC chondrogenesis, thus MeHA-Col II was selected as the optimal injectable hydrogel, and MeHA-Col I/Col II as the preferred bioink. In summary, this study demonstrates how tailoring biomaterial composition and physicochemical properties enables development of pro-chondrogenic hydrogels with potential for minimally invasive delivery to injured articular joints or 3DP of customised regenerative implants for cartilage repair.