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Excessive growth hormone (GH) has been shown to promote joint degeneration in both preclinical and clinical studies. Little is known about the effect of disrupted GH or GH receptor (GHR) on joint health. The goal of this study is to investigate joint pathology in mice with either germline (GHR-/-) or adult inducible (iGHR-/-) GHR deficiency. Knee joints from male and female GHR-/- and WT mice at 24 months of age were processed for histological analysis. Also, knee joints from male and female iGHR-/- and WT mice at 22 months of age were scanned by micro-CT (μCT) for subchondral bone changes and characterized via histology for cartilage degeneration. Joint sections were also stained for the chondrocyte hypertrophy marker, COLX, and the cartilage degeneration marker, ADAMTS-5, using immunohistochemistry. Compared to WT mice, GHR-/- mice had remarkably smooth articular joint surfaces and an even distribution of proteoglycan with no signs of degeneration. Quantitatively, GHR-/- mice had lower OARSI and Mankin scores compared to WT controls. By contrast, iGHR-/- mice were only moderately protected from developing aging-associated OA. iGHR-/- mice had a significantly lower Mankin score compared to WT. However, Mankin scores were not significantly different between iGHR-/- and WT when males and females were analyzed separately. OARSI scores did not differ significantly between WT and iGHR-/- in either individual or combined sex analyses. Both GHR-/- and iGHR-/- mice had fewer COLX+ hypertrophic chondrocytes compared to WT, while no significant difference was observed in ADAMTS-5 staining. Compared to WT, a significantly lower trabecular thickness in the subchondral bone was observed in the iGHR-/- male mice but not in the female mice. However, there were no significant differences between WT and iGHR-/- mice in the bone volume to total tissue volume (BV/TV), bone mineral density (BMD), and trabecular number in either sex. This study identified that both germline and adult-induced GHR deficiency protected mice from developing aging-associated OA with more effective protection in GHR-/- mice.

Objective: This study investigated the protective effects of low fluid shear stress (FSS ≤ 2 dyn/cm²) against interleukin-1β (IL-1β)-induced chondrocyte apoptosis and explored the underlying molecular mechanisms. Methods: Chondrocytes were cultured under four conditions: control, IL-1β stimulation, low FSS, and combined low FSS + IL-1β stimulation. Apoptosis was assessed using Hoechst staining and flow cytometry. Western blotting determined the expression of caspase-3 (CASP3), caspase-8 (CASP8), and NF-κB p65. Quantitative real-time PCR measured miR-143-3p expression. The roles of miR-143-3p and the extracellular signal-regulated kinase 5 (ERK5)/Krüppel-like factor 4 (KLF4) signaling pathway were further investigated using miR-143-3p mimics and inhibitors, an ERK5 inhibitor, and a KLF4 overexpression vector. Results: IL-1β induced significant chondrocyte apoptosis, which was markedly inhibited by low FSS. Mechanistically, low FSS suppressed miR-143-3p expression, thereby enhancing ERK5 signaling. This activated ERK5 subsequently upregulated KLF4 expression, further mitigating IL-1β-induced damage. Importantly, miR-143-3p overexpression under low FSS conditions exacerbated IL-1β-induced apoptosis, while miR-143-3p inhibition attenuated it. Consistent with this, ERK5 inhibition augmented IL-1β-induced apoptosis, whereas KLF4 overexpression suppressed it. Conclusion: Low FSS protects chondrocytes from IL-1β-induced apoptosis by suppressing miR-143-3p and activating the ERK5/KLF4 signaling pathway. This study reveals a novel mechanism by which mechanical stimulation protects cartilage. [ABSTRACT FROM AUTHOR]

BACKGROUND: Long intergenic non-protein coding RNA 00707 (LINC00707) and microRNA-423-5p (miR-423-5p) are both involved in the occurrence and development of osteoarthritis. Starbase predicts that LINC00707 and miR-423-5p have complementary sequences, but whether LINC00707 and miR-423-5p interact to regulate the progress of osteoarthritis still needs further research. OBJECTIVE: To investigate whether LINC00707 targets miR-423-5p to affect chondrocyte injury and inflammatory factor secretion in osteoarthritis. METHODS: Articular chondrocytes were divided into eight groups: (1) blank control group was given no treatment; (2) interleukin (IL)-1β group was cultured with 10 ng/mL IL-1β for 48 hours; (3) IL-1β+si-NC group was transfected with si-NC and then treated with 10 ng/mL IL-1β for 48 hours; (4) IL-1β+si-LINC00707 group was transfected with si-LINC00707 and then treated with 10 ng/mL IL-1β for 48 hours; (5) IL-1β+miR-NC group was transfected with miR-NC and then treated with 10 ng/mL IL-1β for 48 hours; (6) IL-1β+miR-423-5p group was transfected with miR-423-5p mimic and then treated with 10 ng/mL IL-1β for 48 hours; (7) IL-1β+si-LINC00707+anti-miR-NC group was co-transfected with si-LINC00707 and anti-miR-NC and then treated with 10 ng/mL IL-1β for 48 hours; (8) IL-1β+si-LINC00707+anti-miR-423-5p group was co-transfected with si-LINC00707 and anti-miR-423-5p and then treated with 10 ng/mL IL-1β for 48 hours. Relevant tests were performed at the end of the intervention. RESULTS AND CONCLUSION: Compared with the blank control group, the mRNA expression of LINC00707, apoptosis rate, protein expression of C-caspase3 and C-caspase9, and levels of tumor necrosis factor-α and IL-6 in articular chondrocytes were increased in the IL-1β group, while there was a decrease in miR-423-5p expression and IL-10 level (P < 0.05). Compared with the IL-1β group, the mRNA expression of LINC00707, apoptosis rate, protein expression of C-caspase3 and C-caspase9, and levels of tumor necrosis factor-α and IL-6 in articular chondrocytes were decreased in the IL-1β+si-LINC00707 group, while miR-423-5p expression and IL-10 level increased (P < 0.05). Compared with the IL-1β+miR-NC group, the protein expression of C-caspase3 and C-caspase9 and levels of tumor necrosis factor-α and IL-6 in articular chondrocytes were decreased in the IL-1β+miR-423-5p group, while miR-423-5p expression and IL-10 level increased (P < 0.05). Compared with the IL-1β+si-LINC00707+anti-miR-NC group, apoptosis rate, protein expression of C-caspase3 and C-caspase9, and levels of tumor necrosis factor-α and IL-6 in articular chondrocytes were increased in the IL-1β+si-LINC00707+anti-miR-423-5p group, while miR-423-5p expression and IL-10 level decreased (P < 0.05). To conclude, inhibiting LINC00707 by targeting miR-423-5p can reduce IL-1β-induced apoptosis and inflammation in articular chondrocytes. [ABSTRACT FROM AUTHOR]

BACKGROUND: Previous studies have shown that silent mating-type information regulator 2 homolog 1 (SIRT1)-mechanistic target of rapamycin (mTOR) signaling plays an important role in the progression of osteoarthritis. Emodin has a protective effect on osteoarthritic chondrocytes.OBJECTIVE: To investigate the effects of emodin on the proliferation, apoptosis and oxidative stress of osteoarthritic chondrocytes based on the SIRT1-mTOR signaling. METHODS: Rat chondrocytes were isolated and cultured in vitro. Osteoarthritic chondrocyte model in vitro was induced by 10 ng/mL interleukin-1β. Cell counting kit-8 method was used to determine the viability of rat chondrocytes treated with 0, 20, 40, 80, 120, 160 µmol/L emodin, and the appropriate concentration of emodin was selected. Rat chondrocytes isolated and cultured in vitro were randomly divided into control group, model group, low-dose emodin group, high-dose emodin group, EX527 group, and high-dose emodin+EX527 group. In vitro osteoarthritis model was constructed by induction of 10 ng/mL interleukin 1β in all groups except the control group. The cells in the latter four groups were correspondingly treated with emodin or/and EX527. The proliferation and apoptosis of chondrocytes in each group were detected by cell counting kit-8, Edu staining and flow cytometry respectively. The relative content of reactive oxygen species and the levels of malondialdehyde, superoxide dismutase, catalase, and glutathione peroxidase in chondrocytes of rats in each group were measured with the kit. The expression of proteins related to cell matrix degradation, apoptosis and the SIRT1-mTOR pathway-related proteins in each group were detected by western blot. RESULTS AND CONCLUSION: Compared with the control group, the survival rate of chondrocytes, the positive rate of Edu, the levels of superoxide dismutase, catalase, and glutathione peroxidase, and the expression of Bcl-2 and SIRT1 proteins in the model group were decreased, while the apoptosis rate, the relative content of reactive oxygen species, the level of malondialdehyde, the expression of Bax, matrix metalloproteinase 3, matrix metalloproteinase 9 proteins, and p-mTOR/mTOR were increased (P < 0.05). Compared with the model group, the survival rate of chondrocytes, the positive rate of Edu, the levels of superoxide dismutase, catalase, and glutathione peroxidase, and the expression of Bcl-2 and SIRT1 proteins in the low- and high-dose emodin groups were increased, while the apoptosis rate, the relative content of reactive oxygen species, the level of malondialdehyde, the expression of Bax, matrix metalloproteinase 3, matrix metalloproteinase 9 proteins, and p-mTOR/mTOR were decreased (P < 0.05). Compared with the low- and high-dose emodin groups, the indexes of EX527 group showed the opposite trend (P < 0.05). Compared with the high-dose emodin group, the survival rate of chondrocytes, the positive rate of Edu, the levels of superoxide dismutase, catalase, and glutathione peroxidase, and the expression of Bcl-2 and SIRT1 proteins in the high-dose emodin+EX527 group were decreased, while the apoptosis rate, the relative content of reactive oxygen species, the level of malondialdehyde, the expression of Bax, matrix metalloproteinase 3, matrix metalloproteinase 9 proteins, and p-mTOR/mTOR were increased (P < 0.05). To conclude, emodin can inhibit oxidative stress of osteoarthritic chondrocytes by activating the SIRT1-mTOR signaling, thereby promoting chondrocyte proliferation and reducing apoptosis. [ABSTRACT FROM AUTHOR]

In this review, we explore the intricate relationship between glucose metabolism and mechanotransduction pathways, with a specific focus on the role of the Hippo signaling pathway in chondrocyte pathophysiology. Glucose metabolism is a vital element in maintaining proper chondrocyte function, but it has also been implicated in the pathogenesis of osteoarthritis (OA) via the induction of pro-inflammatory signaling pathways and the establishment of an intracellular environment conducive to OA. Alternatively, mechanotransduction pathways such as the Hippo pathway possess the capacity to respond to mechanical stimuli and have an integral role in maintaining chondrocyte homeostasis. However, these mechanotransduction pathways can be dysregulated and potentially contribute to the progression of OA. We discussed how alterations in glucose levels may modulate the Hippo pathway components via a variety of mechanisms. Characterizing the interaction between glucose metabolism and the Hippo pathway highlights the necessity of balancing both metabolic and mechanical signaling to maintain chondrocyte health and optimal functionality. Furthermore, this review demonstrates the scarcity of the literature on the relationship between glucose metabolism and mechanotransduction and provides a summary of current research dedicated to this specific area of study. Ultimately, increased research into this topic may elucidate novel mechanisms and relationships integrating mechanotransduction and glucose metabolism. Through this review we hope to inspire future research into this topic to develop innovative treatments for addressing the clinical challenges of OA. [ABSTRACT FROM AUTHOR]

In the present work, we introduce a scheme for efficient mechanobiological research on the dedifferentiation of chondrocytes (CHs) using a deep neural network (DNN) model that does not require sacrificing the cells and thus, saving resources. Cells are a part of active biological systems subjected to physical stimuli such as mechanical loading. Such loading affects cellular processes including proliferation, differentiation, and the interplay with the surrounding environment. CHs are mechanosensitive cells that produce and maintain the cartilaginous matrix that allows cartilage to bear and distribute mechanical loads in joints; their role in cartilage regeneration and treatment of osteoarthritis (OA) has been the focus of research projects. Nevertheless, it remains an open challenge to fully understand the effect of mechanical stimuli on CHs. One of the unresolved mechanobiological behaviors of CHs is dedifferentiation. Dedifferentiation of CHs is the phenomenon where isolated CHs alter their phenotype when cultured in a 2D in vitro environment over passages. While intact, cobblestone‐shaped CHs mainly produce collagen II, dedifferentiated CHs with elongated shapes produce fibroblastic collagen I. This limits the scalability of a promising treatment for OA, called 'autologous chondrocyte implantation (ACI)'. To overcome this limit, research is being carried out to understand the dedifferentiation mechanism in detail. This proposed scheme is composed of three parts: (i) the cell‐seeded specimens are loaded in a bioreactor system, (ii) an optical microscope is used to obtain the phase‐contrast images of living cells, and (iii) the images are analyzed using a DNN. This scheme provides an efficient tool to analyze the ratio of intact to dedifferentiated CHs while preserving cells on our way to elucidate the effects of mechanical loading on the dedifferentiation of CHs. [ABSTRACT FROM AUTHOR]

In articular cartilage, the pericellular matrix acting as a specialized mechanical microenvironment modulates environmental signals to chondrocytes through mechanotransduction. Matrix viscoelastic alterations during cartilage development and osteoarthritis (OA) degeneration play an important role in regulating chondrocyte fate and cartilage matrix homeostasis. In recent years, scientists are gradually realizing the importance of matrix viscoelasticity in regulating chondrocyte function and phenotype. Notably, this is an emerging field, and this review summarizes the existing literatures to the best of our knowledge. This review provides an overview of the viscoelastic properties of hydrogels and the role of matrix viscoelasticity in directing chondrocyte behavior. In this review, we elaborated the mechanotransuction mechanisms by which cells sense and respond to the viscoelastic environment and also discussed the underlying signaling pathways. Moreover, emerging insights into the role of matrix viscoelasticity in regulating chondrocyte function and cartilage formation shed light into designing cell‐instructive biomaterial. We also describe the potential use of viscoelastic biomaterials in cartilage tissue engineering and regenerative medicine. Future perspectives on mechanobiological comprehension of the viscoelastic behaviors involved in tissue homeostasis, cellular responses, and biomaterial design are highlighted. Finally, this review also highlights recent strategies utilizing viscoelastic hydrogels for designing cartilage‐on‐a‐chip. Summary: It is well known that viscoelasticity has been found to be a universal characteristic of living tissues, cells, and molecules.Matrix viscoelasticity, as a critical physical cues of cartilage matrix microenvironment, regulates collective chondrocyte fate and cartilage formation. But, how matrix viscoelasticity affects the chondrocyte behavior and function remains unclear.Growing evidence has demonstrated that matrix viscoelasticity regulates chondrocytes in some ways not anticipated from previous mechanisms of mechanotransduction, which was based on purely substrate stiffness.In this review, we introduce recent work elucidating the effect of matrix viscoelasticity on chondrocytes morphology, phenotype, function, inflammatory response, and signaling pathways.These findings have provided insights into viscoelasticity as a design parameter for cartilage tissue engineering, regenerative medicine, and cartilage‐on‐a‐chip, even shed light into the pathological mechanism of joint growth and degeneration. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to investigate whether flow fluid shear stress (FFSS)‐mediated signal transduction affects the function of Piezo1 ion channel in chondrocyte and to further explore the role of mechanical overloading in development of temporomandibular joint osteoarthritis (TMJ OA). Methods: Immunohistochemical staining was used to determine the expression of Piezo1 in TMJ OA tissue collected from rat unilateral anterior crossbite (UAC) models. Chondrocytes harvested from normal adult SD rats were treated with FFSS (0, 4, 8, 12 dyn/cm2) in vitro. Immunofluorescent staining, real‐time polymerase chain reaction, western blotting, flow cytometry and phalloidin assay were performed to detect the changes of cellular morphology as well as the expression of Piezo1 and certain pro‐inflammatory and degradative factors in chondrocyte. Results: Immunohistochemical analysis revealed that significantly increased Piezo1 expression was associated with UAC stimulation (p <.05). As applied FFSS escalated (4, 8 and 12 dyn/cm2), the expression levels of Piezo1, ADAMTS‐5, MMP‐13 and Col‐X gradually increased, compared with the non‐FFSS group (p <.05). Administering Piezo1 ion channel inhibitor to chondrocytes beforehand, it was observed that expression of ADAMTS‐5, MMP‐13 and Col‐X was substantially decreased following FFSS treatment (p <.05) and the effect of cytoskeletal thinning was counteracted. The activated Piezo1 ion channel enhanced intracellular Ca2+ excess in chondrocytes during abnormal mechanical stimulation and the increased intracellular Ca2+ thinned the cytoskeleton of F‐actin. Conclusions: Mechanical overloading activates Piezo1 ion channel to promote pro‐inflammation and degradation and to increase Ca2+ concentration in chondrocyte, which may eventually result in TMJ OA. [ABSTRACT FROM AUTHOR]

Tumor necrosis factor alpha (TNFα), as a key pro-inflammatory cytokine, plays a central role in joint diseases. In recent years, numerous models of TNFα-induced cartilage inflammation have been developed. However, due to the significant differences between these models and the lack of consensus in their construction, it becomes difficult to compare the results of different studies. Therefore, we summarized and compared these models based on important parameters for model construction, such as cell source, cytokine concentration, stimulation time, mechanical stimulation, and more. We attempted to analyze the advantages and disadvantages of each model and provide a compilation of the analytical methods used in previous studies. Currently, TNFα chondrocyte inflammation models can be categorized into four main types: monolayer-based, construct-based, explant-based TNFα chondrocyte inflammation models, and miscellaneous TNFα chondrocyte inflammation models. The most commonly used models were the monolayer-based TNFα chondrocyte inflammation models (42.86% of cases), with 10 ng/mL TNFα being the most frequently used concentration. The most frequently used chondrocyte cell passage is passage 1 (50%). Human tissues were most frequently used in experiments (51.43%). Only five articles included models with mechanical stimulations. We observed variations in design conditions between different models. This systematic review provides the essential experimental characteristics of the available chondrocyte inflammation models with TNFα, and it provides a platform for better comparison between existing and new studies in this field. It is essential to perform further experiments to standardize each model and to find the most appropriate experimental parameters. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a chronic joint disease with an unclear cause characterized by secondary osteophytes and degenerative changes in the articular cartilage. More than 250 million people are expected to be affected by it by 2050, putting a tremendous socioeconomic strain on the entire world. OA cannot currently be treated with any effective medications that change the illness. Over time, chondrocytes undergo gradual metabolic, structural, and functional changes as a result of aging or abuse. The degenerative progression of osteoarthritis is significantly influenced by the imbalance of chondrocyte homeostasis. By continuously recycling and rebuilding macromolecules or organelles, autophagy functions as a crucial regulatory system to maintain homeostasis during an individual's growth and development. This review uses chondrocytes as its starting point and establishes a strong connection between autophagy and osteoarthritis in order to thoroughly examine the mechanisms behind chondrocyte autophagy in osteoarthritis. Biomarkers of chondrocyte autophagy will be identified, and prospective targeted medications and novel treatment approaches for slowing or preventing the course of OA will be developed based on chondrocyte senescence, autophagy, and apoptosis in OA. Key messages: Currently, OA has not been treated with any drugs that can effectively cure it. We hope that by exploring specific targets in the course of osteoarthritis, we can promote the progress of treatment strategies. The degenerative progression of osteoarthritis is significantly influenced by the imbalance of chondrocyte balance. Through the continuous recovery and reconstruction of macromolecules or organelles, autophagy is an important regulatory system for maintaining homeostasis during individual growth and development. In this paper, the close relationship between autophagy and osteoarthritis was established with chondrocytes as the starting point, in order to further explore the mechanism of chondrocyte autophagy in osteoarthritis. The development process of osteoarthritis was studied from the perspective of chondrocytes, and the change of autophagy level had a significant impact on osteoarthritis. Chondrocyte autophagy is mainly determined by intracellular mitochondrial autophagy, so we are committed to finding relevant molecules. Through PI3K/AKT- and MAPK-related pathways, the biomarkers of chondrocyte autophagy were identified, and chondrocyte senescence, autophagy, and apoptosis based on osteoarthritis provided a constructive idea for the development of prospective targeted drugs and new therapies to slow down or prevent the progression of osteoarthritis. [ABSTRACT FROM AUTHOR]

BACKGROUND: Upregulation of hedgehog protein signaling can increase the expression of osteoarthritis markers, Runx2, a disintegrin and metalloproteinase with thrombospondin motifs, collagen type X alpha 1, and matrix metalloproteinase 13, while inhibition of hedgehog proteins attenuates the severity of osteoarthritis. It is speculated that osteoarthritic chondrocytes can influence bone formation by affecting osteoblasts through the Indian hedgehog protein (IHH) signaling pathway. OBJECTIVE: To investigate the effect of human osteoarthritic chondrocytes on subchondral osteoblasts. METHODS: Tibial plateau specimens from patients with osteoarthritis were collected. Chondrocytes were extracted using enzymatic digestion, and osteoblasts were extracted using enzymatic pre-digestion + bone block method. Chondrocytes were identified by toluidine blue staining and immunofluorescence and osteoblasts were identified by alkaline phosphatase staining and immunofluorescence. Chondrocytes were cultured in sodium alginate beads to maintain chondrocyte phenotype and co-cultured with osteoblasts. The co-culture system was added with IHH signaling pathway inhibitor (cyclopamine, 10 nmol/L) and activator (purmorphamine, 10 nmol/L) separately. After 48 hours of co-culture, osteoblasts from each group were collected, mRNA expressions of Gli1, osteoprotegerin, Runx2, parathyroid hormone-related peptide, alkaline phosphatase, receptor activator of nuclear factor-kB ligand (RANKL) and osteocalcin were detected by qRT-PCR, and protein expressions of GLi1, oseoprotegerin and RANKL in osteoblasts were detected by western blot. RESULTS AND CONCLUSION: The mRNA expression levels of GLi1, osteoprotegerin and RUNX2 in osteoblasts were significantly increased, while the mRNA expression levels of parathyroid hormone-related peptide were decreased (P < 0.05) when co-cultured with human osteoarthritic chondrocytes. The mRNA and protein levels of Gli1 were significantly decreased after the addition of IHH signaling pathway inhibitor (cyclopamine) (P < 0.05), and the mRNA and protein levels of Gli1 were significantly increased after the addition of IHH signaling pathway activator (purmorphamine) (P < 0.05). Osteoprotegerin showed the same trend as Gli1 in the experiment. The osteoprotegerin/RANKL ratio followed the same trend as osteoprotegerin. To conclude, human osteoarthritic chondrocytes can promote the expression of Gli1, osteoprotegerin, Runx2 and other proteins in osteoblasts. The upregulation of osteoprotegerin is related to the IHH signaling pathway. Osteoarthritic chondrocytes can up-regulate the expression of osteoprotegerin in osteoblasts through the IHH signaling pathway and thus up-regulate the osteoprotegerin/RANKL ratio, which will contribute to bone formation in subchondral bone. [ABSTRACT FROM AUTHOR]

BACKGROUND: Studies have shown that hesperetin exerts protective effects on chondrocytes through a variety of mechanisms or signaling pathways, but the protective mechanisms have not been fully elucidated. OBJECTIVE: To investigate the effect of hesperetin on lipopolysaccharide-induced inflammatory degeneration of chondrocytes. METHODS: Knee joint chondrocytes from suckling Sprague-Dawley rats were isolated and cultured in vitro, and identified by Safranine O staining. The cytotoxicity of hesperetin on chondrocytes was determined by the MTT assay to ensure the optimal concentration of hesperetin. Chondrocytes were randomly divided into three groups, including the control group, model group, and experimental group. The cellular model of osteoarthritis was established in the latter two groups by simulating chondrocytes with lipopolysaccharide. Chondrocytes in the experimental group was then intervened with hesperetin for 24 hours. Calcein-AM/EthD-I staining was used to detect cell viability. Immunohistochemical staining was performed to determine the expression of type II collagen in chondrocytes. Intracellular reactive oxygen species level was detected by a reactive oxygen species detection kit. Total glutathione level was detected by ELISA. Real-time fluorescent PCR was employed to detect the expression of interleukin 1β, interleukin 6, tumor necrosis factor α and type II collagen. RESULTS AND CONCLUSION: Safranine O staining results showed that the cells extracted were chondrocytes. Cytotoxicity test results showed 0.5 μmol/L hesperetin had the most significant effect on chondrocyte vitality. Compared with the control group, the model group showed a decrease in chondrocyte proliferation ability, an increase in reactive oxygen species levels, a decrease in total glutathione levels, an increase in type II collagen degradation, an increase in the levels of interleukin 1β, interleukin 6, and tumor necrosis factor α, and a decrease in the expression of type II collagen (P < 0.05). Compared with the model group, in the experimental group, the proliferation ability of chondrocytes increased, reactive oxygen species levels decreased, total glutathione levels increased, and type II collagen degradation decreased, levels of interleukin 1β, interleukin 6, and tumor necrosis factor α downregulated, and the expression of type II collagen upregulated (P < 0.05). To conclude, hesperetin has a protective effect on lipopolysaccharide-induced inflammatory degeneration of osteoarthritic chondrocytes, and the mechanism may be associated with inhibition of reactive oxygen species-mediated oxidative stress. [ABSTRACT FROM AUTHOR]

Oxytocin (OT) is a posterior pituitary hormone that, in addition to its role in regulating childbirth and lactation, also exerts direct regulatory effects on the skeleton through peripheral OT and oxytocin receptor (OTR). Bone marrow mesenchymal stem cells (BMSCs), osteoblasts (OB), osteoclasts (OC), chondrocytes, and adipocytes all express OT and OTR. OT upregulates RUNX2, BMP2, ALP, and OCN, thereby enhancing the activity of BMSCs and promoting their differentiation towards OB rather than adipocytes. OT also directly regulates OPG/RANKL to inhibit adipocyte generation, increase the expression of SOX9 and COMP, and enhance chondrocyte differentiation. OB can secrete OT, exerting influence on the surrounding environment through autocrine and paracrine mechanisms. OT directly increases OC formation through the NκB/MAP kinase signaling pathway, inhibits osteoclast proliferation by triggering cytoplasmic Ca2+ release and nitric oxide synthesis, and has a dual regulatory effect on OCs. Under the stimulation of estrogen, OB synthesizes OT, amplifying the biological effects of estrogen and OT. Mediated by estrogen, the OT/OTR forms a feedforward loop with OB. Apart from estrogen, OT also interacts with arginine vasopressin (AVP), prostaglandins (PGE2), leptin, and adiponectin to regulate bone metabolism. This review summarizes recent research on the regulation of bone metabolism by OT and OTR, aiming to provide insights into their clinical applications and further research. [ABSTRACT FROM AUTHOR]

Low back pain (LBP) is largely attributed to intervertebral disc degeneration (IVDD), of which the endplate changes are an important component. However, the alterations in cell fate and properties within the endplates during degeneration remain unknown. Here, we firstly performed the single-cell RNA-sequencing analysis (scRNA-seq) of the cells focusing on degenerative human endplates. By unsupervised clustering of the 8,534 single-cell based on the gene expression, we identified nine distinct cell types. We employed Gene ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analysis, and the single-cell regulatory network inference and clustering (SCENIC) to determine the enriched pathways and transcriptional activities across seven chondrocyte subpopulations. Furthermore, two cell fates of chondrocyte differentiation were found by trajectory analysis, one was enriched in inflammation-related genes, and the other was related to extracellular matrix (ECM). Additionally, the intercellular interactions of macrophages (MA) and chondrocytes, T cells/natural killer cells (T/NK) and chondrocytes were examined by ligand-receptor pairs analysis, showing the important regulative function of FN1 from MA and CD74 from T/NK during endplate degeneration. Overall, our findings provide novel perspectives on the endplate degeneration at the single-cell level and a whole-transcriptome size. [ABSTRACT FROM AUTHOR]

Articular cartilage damage and wear can result in cartilage degeneration, ultimately culminating in osteoarthritis. Current surgical interventions offer limited capacity for cartilage tissue regeneration and offer only temporary alleviation of symptoms. Tissue engineering strategies are increasingly recognized as promising modalities for cartilage restoration. Currently, various biological scaffolds utilizing tissue engineering materials are extensively employed in both fundamental and clinical investigations of cartilage repair. In order to optimize the cartilage repair ability of tissue engineering scaffolds, researchers not only optimize the structure and properties of scaffolds from the perspective of materials science and manufacturing technology to enhance their histocompatibility, but also adopt strategies such as loading cells, cytokines, and drugs to promote cartilage formation. This review provides an overview of contemporary tissue engineering strategies employed in cartilage repair, as well as a synthesis of existing preclinical and clinical research. Furthermore, the obstacles faced in the translation of tissue engineering strategies to clinical practice are discussed, offering valuable guidance for researchers seeking to address these challenges. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is the most prevalent cartilage degenerative and low-grade inflammatory disease of the whole joint. However, there are currently no FDAapproved drugs or global regulatory agency-approved treatments OA disease modification. Therefore, it’s essential to explore novel effective therapeutic strategies for OA. In our study, we investigated the effects of AFK-PD, a novel pyridone agent, on the development of OA induced by destabilization of the medial meniscus (DMM) in vivo, and its impact on the function of chondrocytes treated with IL-1β in vitro. Our results demonstrated AFK-PD alleviated OA progression through inhibiting cartilage degeneration, articular inflammation and osteophyte formation. Notably, AFK-PD inhibited chondrocyte inflammation and synovial macrophage M1 polarization, leading to the attenuation of articular inflammation. Additionally, AFK-PD promoted chondrocyte anabolism while mitigating catabolism and apoptosis, effectively inhibiting cartilage degeneration. Mechanistically, AFK-PD suppressed the expression of key signaling molecules involved in the MAPK pathway, such as p-ERK1/2 and p-JNK, as well as the NF-κB signaling molecule p-p65, in IL-1βinduced chondrocytes. These findings suggest AFK-PD ameliorates the development of OA by protecting chondrocyte functions and inhibiting articular inflammation in chondrocytes and synovial macrophages. Overall, our study highlights AFK-PD as a promising therapeutic candidate for the treatment of OA. [ABSTRACT FROM AUTHOR]

BACKGROUND: Heterotopic ossification of skeletal muscle is a clinically serious complication. For heterotopic ossification of skeletal muscles, the cells involved in the process of heterotopic ossification remain unclear. OBJECTIVE: To investigate the involvement of myocytes, fascia cells, and endothelial cells in the process of heterotopic ossification in skeletal muscle and to observe the cell origin of heterotopic ossification in skeletal muscle induced by bone morphogenetic protein 4. METHODS: Both C2C12 cells and the myotubes formed by the C2C12 cells in the induction medium were cultured, and 500 ng/mL bone morphogenetic protein 4 was added to the medium respectively, and whether the C2C12 cells and myotubes continued to proliferate within 10 days under the treatment were observed under a microscope. Myogenic cells (L6, derived from rats) and fibroblast-derived cells (derived from human) were co-cultured. After treatment with 500 ng/mL bone morphogenetic protein 4 and 10 ng/mL transforming growth factor-β, osteogenic and chondrogenic differentiation potential within 21 days were observed using Safranine O staining and Alcian blue staining. Using transgenic animal FVB/N-TgN (TIE2-LacZ) 182Sato mice, 15 μL of adeno-associated virus-bone morphogenetic protein 4 (5 × 1010 PFU/mL) were implanted in the thigh muscle space of genetic mice for 10 and 14 days. X-gal staining was used to observe the formation of new blood vessel endothelium in the differentiated bone. RESULTS AND CONCLUSION: (1) Bone morphogenetic protein 4 caused myotube breakdown and increased C2C12 cell proliferation. Compared with other groups, the pure fibroblast-derived cell group had a higher area of positive alcian blue and safarin O staining (P < 0.05) and a lower area of alkaline phosphatase staining (P < 0.05), while the pure L6 group had a bigger area of alkaline phosphatase staining (P < 0.05) but a smaller area of positive alcian blue and safarin O staining (P < 0.05). (2) Transplantation of adeno-associated virus-bone morphogenetic protein 4-adsorbed gelatin sponge into FVB/N-TgN (TIE2-LacZ)182Sato mice resulted in heterotopic ossification. (3) X-gal staining results demonstrated that there was no obvious staining in chondrocytes and differentiated bones and Tie2+ endothelial cells did not participate in the formation of the alienated bone. (4) These findings verify that fibroblasts are the primary source of osteoblasts during the adeno-associated virus-bone morphogenetic protein 4-induced ectopic endochondral ossification in skeletal muscle, but myogenic cells are the main source of osteoblasts. Tie2+ endothelial cells might not be the cell source for cartilage and bone. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of silencing miR-126 on proliferation, apoptosis, and inflammatory response of osteoarticular chondrocytes and their mechanism. Methods Human osteoarticular chondrocytes HC-a were cultured in vitro. HC-a cells in the third generation, which were in the logarithmic growth phase and had good growth status, were randomly divided into the control group, IL-1β group, IL-1β + miR-NC group, and IL-1β + miR-126 inhibitor group, respectively. Cells in the IL-1β + miR-NC group and IL-1β + miR-126 inhibitor group were transfected with miR-NC and miR-126 inhibitor, respectively. Except for the control group, the other three groups were treated with 10 ng/mL IL-1β induction to construct the cell models of osteoarthritis (OA) . The cells of each group were collected, and the expression of miR-126 was detected by RT-qPCR, cell proliferation activity was detected by CCK-8, apoptosis rate was detected by flow cytometry, and inflammatory factors TNF-α, IL-6 and IFN-γ in the culture supernatant were detected by ELISA. The expression levels of SIRT1 and Caspase-3 proteins were detected by Western blotting. Results Compared with the control group, the relative expression of miR-126 increased, the cell proliferation activity decreased, apoptosis rate increased, the levels of TNF-α, IL-6 and IFN-γ in culture supernatance increased, the relative expression of SIRT1 protein decreased, and the relative expression of Caspase-3 protein increased in the IL-1β group (all P<0. 05) . Compared with the IL-1β + miR-NC group, the relative expression decreased, cell proliferation activity increased, apoptosis rate decreased, the levels of TNF-α, IL-6 and IFN-γ in culture supernatant decreased, the relative expression of SIRT1 protein increased, and the relative expression of Caspase-3 protein decreased in the miR-126 inhibitor group (all P<0. 05) . There were no significant differences in the above indexes between IL-1β group and IL-1β + miR-NC group (all P>0. 05) . Conclusion Silencing miR-126 can promote the proliferation of OA cells and inhibit apoptosis and inflammation, which may be related to the regulation of SIRT1 protein expression. [ABSTRACT FROM AUTHOR]

Background: Chondrocyte viability is associated with the clinical success of osteochondral allograft (OCA) transplantation. Purpose: To investigate the effect of distal femoral OCA plug harvest and recipient site preparation on regional cell viability using traditional handheld saline irrigation versus saline submersion. Study Design: Controlled laboratory study. Methods: For each of 13 femoral hemicondyles, 4 cartilage samples were harvested: (1) 5-mm control cartilage, (2) 15-mm OCA donor plug harvested with a powered coring reamer and concurrent handheld saline irrigation ("traditional"), (3) 15-mm OCA donor plug harvested while submerged under normal saline ("submerged"), and (4) 5-mm cartilage from the peripheral rim of a recipient socket created with a 15-mm cannulated counterbore reamer to a total depth of 7 mm with concurrent handheld saline irrigation ("recipient"). The 15 mm–diameter plugs were divided into the central 5 mm and the peripheral 5 mm (2 edges) for comparisons. Samples were stained using calcein and ethidium, and live/dead cell percentages were calculated and compared across groups. Results: Compared with the submerged group, the traditional group had significantly lower percentages of live cells across the whole plug (71.54% ± 4.82% vs 61.42% ± 4.98%, respectively; P =.003), at the center of the plug (72.76% ± 5.87% vs 62.30% ± 6.11%, respectively; P =.005), and at the periphery of the plug (70.93% ± 4.51% vs 60.91% ± 4.75%, respectively; P =.003). The traditional group had significantly fewer live cells in all plug regions compared with the control group (77.51% ± 9.23%; P <.0001). There were no significant differences in cell viability between the control and submerged groups (whole: P =.590; center: P =.713; periphery: P =.799). There were no differences between the central and peripheral 5-mm plug regions for the traditional (62.30% ± 6.11% vs 60.91% ± 4.75%, respectively; P =.108) and submerged (72.76% ± 5.87% vs 70.93% ± 4.51%, respectively; P =.061) groups. The recipient group (61.10% ± 5.02%) had significantly lower cell viability compared with the control group (P <.0001) and the periphery of the submerged group (P =.009) but was equivalent to the periphery of the traditional group (P =.990). Conclusion: There was a significant amount of chondrocyte death induced by OCA donor plug harvesting using a powered coring reamer with traditional handheld saline irrigation, which was mitigated by harvesting the plug while the allograft was submerged under saline. Clinical Relevance: Mitigating this thermally induced damage by harvesting the OCA plug while the allograft was submerged in saline maintained chondrocyte viability throughout the plug and may help to improve the integration and survival of OCAs. [ABSTRACT FROM AUTHOR]

Introduction: Osteoarthritis is a degenerative condition of the cartilage, often common among the population and occurs frequently with aging. Many factors are decisive for the development of its pathogenesis such as age, obesity, trauma, mechanical load, and modification of synovial biology. The main features of osteoarthritis are chondrocytes and cartilage matrix loss, which lead to pain, loss of function of the whole joint, and disability, representing a relevant health problem. Recently, a new therapeutic approach based on cell therapy has been studying the regenerative ability of mesenchymal stem cells for osteoarthritic chondrocytes. Aim: This in vitro study clarifies the regenerative effects of multipotent adipose-derived stem cells and the pluripotent amniotic epithelial stem cells on arthrosis chondrocytes by performing co-culture experiments. Methods: We studied the regenerative potential of secretome (soluble factors and extracellular vesicles), mesenchymal stem cells, and the adipose stromal vascular fraction. The regenerative effects were evaluated by gene and protein expression analysis of articular cartilage-specific genes and proteins like col2a1, acan, and sox9. Results: Mesenchymal stem cells, secretome, and adipose stromal vascular fractions influenced the cartilage genes and protein expression. Conclusions: The results indicate that the treatment with mesenchymal stem cells could be the best biological approach for cartilage regenerative medicine. [ABSTRACT FROM AUTHOR]

Desbuquois dysplasia type 1 (DBQD1) is a recessive chondrodysplasia caused by mutations in the CANT1 gene, encoding for the Golgi Calcium-Activated Nucleotidase 1 (CANT1). The enzyme hydrolyzes UDP, the by-product of glycosyltransferase reactions, but it might play other roles in different cell types. Using a Cant1 knock-out mouse, we demonstrated that CANT1 is crucial for glycosaminoglycan (GAG) synthesis; however, its impact on the biochemical properties of cartilage proteoglycans remains unknown. Thus, in this work, we characterized decorin and aggrecan from primary chondrocyte cultures and cartilage biopsies of mutant mice at post-natal day 4 by Western blots and further investigated their distribution in the cartilage extracellular matrix (ECM) by immunohistochemistry. We demonstrated that the GAG synthesis defect caused by CANT1 impairment led to the synthesis and secretion of proteoglycans with shorter GAG chains compared with wild-type animals. However, this alteration did not result in the synthesis and secretion of decorin and aggrecan in the unglycanated form. Interestingly, the defect was not cartilage-specific since also skin decorin showed a reduced hydrodynamic size. Finally, immunohistochemical studies in epiphyseal sections of mutant mice demonstrated that the proteoglycan structural defect moderately affected decorin distribution in the ECM. [ABSTRACT FROM AUTHOR]

Nutritional epigenomics is exceptionally important because it describes the complex interactions among food compounds and epigenome modifications. Phytonutrients or bioactive compounds, which are secondary metabolites of plants, can protect against osteoarthritis by suppressing the expression of inflammatory and catabolic mediators, modulating epigenetic changes in DNA methylation, and the histone or chromatin remodelling of key inflammatory genes and noncoding RNAs. The combination of natural epigenetic modulators is crucial because of their additive and synergistic effects, safety and therapeutic efficacy, and lower adverse effects than conventional pharmacology in the treatment of osteoarthritis. In this review, we have summarized the chondroprotective properties of bioactive compounds used for the management, treatment, or prevention of osteoarthritis in both human and animal studies. However, further research is needed into bioactive compounds used as epigenetic modulators in osteoarthritis, in order to determine their potential value for future clinical applications in osteoarthritic patients as well as their relation with the genomic and nutritional environment, in order to personalize food and nutrition together with disease prevention. [ABSTRACT FROM AUTHOR]

Inflammation models are widely used in the in vitro investigation of new therapeutic approaches for osteoarthritis. TNFα (tumor necrosis factor alpha) plays an important role in the inflammatory process. Current inflammation models lack uniformity and make comparisons difficult. Therefore, this study aimed to systematically investigate whether the effects of TNFα are concentration-dependent and whether chondrocyte expansion has an effect on the inflammatory model. Bovine chondrocytes were enzymatically isolated, expanded to passages 1–3, and transferred into a 3D pellet culture. Chondrocyte pellets were stimulated with recombinant bovine TNFα at different concentrations for 48 h to induce inflammation. Gene expression of anabolic (collagen 2, aggrecan, cartilage oligomeric protein (COMP)), catabolic (matrix metalloproteinases (MMP3, MMP13)), dedifferentiation (collagen 1) markers, inflammation markers (interleukin-6 (IL-6), nuclear factor kappa B (NFkB), cyclooxygenase-2 (COX), prostaglandin-E-synthase-2 (PTGES2)), and the apoptosis marker caspase 3 was determined. At the protein level, concentrations of IL-6, nitric oxide (NO), and sulfated glycosaminoglycans (GAG) were evaluated. Statistical analysis was performed using the independent t-test, and significance was defined as p < 0.05. In general, TNFα caused a decrease in anabolic markers and an increase in the expression of catabolic and inflammatory markers. There was a concentration-dependent threshold of 10 ng/mL to induce significant inflammatory effects. Most of the markers analyzed showed TNFα concentration-dependent effects (COMP, PRG4, AGN, Col1, MMP3, and NFkB). There was a statistical influence of selected gene expression markers from different passages on the TNFα chondrocyte inflammation model, including Col2, MMP13, IL-6, NFkB, COX2, and PTGES2. Considering the expression of collagen 2 and MMP3, passage 3 chondrocytes showed a higher sensitivity to TNFα stimulation compared to passages 1 and 2. On the other hand, MMP13, IL-6, NFkB, and caspase 3 gene expression were lower in P3 chondrocytes compared to the other passages. On the protein level, inflammatory effects showed a similar pattern, with cytokine effects starting at 10 ng/mL and differences between the passages. TNFα had a detrimental effect on cartilage, with a clear threshold observed at 10 ng/mL. Although TNFα effects showed concentration-dependent patterns, this was not consistent for all markers. The selected passage showed a clear influence, especially on inflammation markers. Further experiments were warranted to explore the effects of TNFα concentration and passage in long-term stimulation. [ABSTRACT FROM AUTHOR]

Chondrocyte hypertrophic differentiation is a main event leading to articular cartilage degradation in osteoarthritis. It is associated with matrix remodeling and mineralization, the dynamics of which is not well characterized during chondrocyte hypertrophic differentiation in articular cartilage. Based on an in vitro model of progressive differentiation of immature murine articular chondrocytes (iMACs) into prehypertrophic (Prehyp) and hypertrophic (Hyp) chondrocytes, we performed kinetics of chondrocyte differentiation from Prehyp to Hyp to follow matrix mineralization and remodeling by immunofluorescence, biochemical, molecular, and physicochemical approaches, including atomic force microscopy, scanning electron microscopy associated with energy-dispersive X-ray spectroscopy (SEM–EDS), attenuated total reflection infrared analyses, and X-ray diffraction. Chondrocyte apoptosis was determined by TUNEL assay. The results show the formation of a mineral phase 7 days after Hyp induction, which spreads within the matrices to form poorly crystalline carbonate-substituted hydroxyapatite after 14 days, then the proportions of crystalline relative to amorphous content increases over time. Hyp differentiation also induced a matrix turnover that occurs over the first 7 days, characterized by a decrease in type II collagen and aggrecan and the concomitant appearance of type X collagen. This is accompanied by an increase in the enzymatic activity of MMP-13, the main collagenase in cartilage. The number of apoptotic chondrocytes slightly increased with Hyp differentiation and SEM–EDS analyses detected phosphorus-rich structures that could correspond to apoptotic bodies. Our findings highlight the mechanisms of matrix remodeling events leading to the mineralization of articular cartilage that may occur in osteoarthritis. [ABSTRACT FROM AUTHOR]

The objective of the present study was to review the molecular mechanisms of the adverse effects of environmental pollutants on chondrocytes and extracellular matrix (ECM). Existing data demonstrate that both heavy metals, including cadmium (Cd), lead (Pb), and arsenic (As), as well as organic pollutants, including polychlorinated dioxins and furans (PCDD/Fs) and polychlorinated biphenyls (PCB), bisphenol A, phthalates, polycyclic aromatic hydrocarbons (PAH), pesticides, and certain other organic pollutants that target cartilage ontogeny and functioning. Overall, environmental pollutants reduce chondrocyte viability through the induction apoptosis, senescence, and inflammatory response, resulting in cell death and impaired ECM production. The effects of organic pollutants on chondrocyte development and viability were shown to be mediated by binding to the aryl hydrocarbon receptor (AhR) signaling and modulation of non-coding RNA expression. Adverse effects of pollutant exposures were observed in articular and growth plate chondrocytes. These mechanisms also damage chondrocyte precursors and subsequently hinder cartilage development. In addition, pollutant exposure was shown to impair chondrogenesis by inhibiting the expression of Sox9 and other regulators. Along with altered Runx2 signaling, these effects also contribute to impaired chondrocyte hypertrophy and chondrocyte-to-osteoblast trans-differentiation, resulting in altered endochondral ossification. Several organic pollutants including PCDD/Fs, PCBs and PAHs, were shown to induce transgenerational adverse effects on cartilage development and the resulting skeletal deformities. Despite of epidemiological evidence linking human environmental pollutant exposure to osteoarthritis or other cartilage pathologies, the data on the molecular mechanisms of adverse effects of environmental pollutant exposure on cartilage tissue were obtained from studies in laboratory rodents, fish, or cell cultures and should be carefully extrapolated to humans, although they clearly demonstrate that cartilage should be considered a putative target for environmental pollutant toxicity. [ABSTRACT FROM AUTHOR]

Objective: Chondrocyte apoptosis has been considered a crucial mechanism that is responsible for cartilage destruction in osteoarthritis (OA). The mechanism of interleukin-37 (IL-37) on chondrocyte apoptosis has not been clearly determined in the pathogenesis of OA. Here, we explored the role of IL-37 in the regulation of cellular apoptosis in rat chondrocytes stimulated by IL-1β. Methods: Rat chondrocytes were used in in vitro study, and were stimulated with IL-1β (10 ng/mL) and/or recombinant IL-37 (rIL-37; 100 ng/mL) after cytotoxicity assessments using these cytokines were conducted. After rIL-37 treatment of chondrocytes stimulated with IL-1β, the cell proliferation assay, apoptosis assays, including expression of mitochondrial apoptosis-related markers, flow cytometry analysis of annexin V-FITC/propidium iodide (PI), cell cycle analysis, and Hoechst 33342 staining, and reactive oxygen species (ROS) measurement were used. Results: IL-1β induced expression of inflammatory cytokines and triggered degradation of the extracellular matrix of rat chondrocytes, but this effect was significantly attenuated by rIL-37 treatment. Enhanced ROS generation following IL-1β stimulation was reduced in a dose-dependent manner after stimulation with rIL-37. IL-1β induced pro-apoptotic markers and suppressed anti-apoptotic markers in rat chondrocytes. Flow cytometry using annexin V-FITC/PI revealed that IL-1β increased the apoptosis rate of rat chondrocytes, and that this effect was markedly reversed by treatment with rIL-37. Conclusions: IL-37 potently attenuated IL-1β-mediated apoptosis of rat chondrocytes by blocking ROS production. This study suggests that IL-37 can serve as a novel anti-cytokine therapy in OA by blocking chondrocyte apoptosis. [ABSTRACT FROM AUTHOR]

Articular cartilage lesions are challenging to regenerate, prompting the investigation of novel biomaterial-based therapeutic approaches. Extracellular matrix (ECM)-derived biomaterials are a promising option for this purpose; however, to date, the combination of amniotic membrane (AMM) and articular cartilage (ACM) has not been tested. This study evaluated different concentrations of soluble extracts from the decellularized ECM of amniotic membrane (dAMM) and articular cartilage (dACM), both individually and in combination, to determine their ability to maintain the chondrogenic phenotype in human chondrocytes. After the decellularization process 90–99% of the cellular components were removed, it retains nearly 100% of type 2 collagen and 70% of aggrecan (ACAN) for dACM, and approximately 90% of type IV collagen and 75% of ACAN for dAMM. The biological activity of soluble extracts from dACM and dAMM were evaluated on human chondrocytes. After 72 h, 1.5 mg/mL of dACM and 6 mg/mL of dAMM significantly increased (p < 0.05) the proliferation and expression of SOX9 and ACAN. Also, the combination of both (1.5 mg/mL dACM and 6 mg/mL dAMM) showed synergistic effects, enhancing chondrocyte proliferation, maintaining chondrogenic lineage, and increasing the production of cartilage ECM components, such as COLII (1.5-fold), SOX9 (2-fold), and ACAN (2-fold). These results suggest that the combined use of dACM and dAMM has potential for cartilage regeneration. [ABSTRACT FROM AUTHOR]

BACKGROUND: The Yi people have used Sambucus adnata Wall to treat fractures, bruises, arthritis, etc. The author’s group found that the active site aqueous extract (SAW-A) and dichloromethane extract (SAW-B) can promote fracture healing by promoting the expression of genes related to osteoblast proliferation, differentiation and maturation, differentiation and maturation. However, the therapeutic effect of methanol extract (SAW-C) at its active site on osteoarthritis is unclear. OBJECTIVE: To investigate the therapeutic effect of Sambucus adnata Wall extract on osteoarthritis, and to identify the effective targets of Sambucus adnata Wall extract in the treatment of osteoarthritis by network pharmacology technology. METHODS: A rat osteoarthritis model was replicated by anterior cruciate ligamentectomy and model rats were then treated with methanol extract of Sambucus adnata Wall by gavage for 21 days. On the 21st day after modeling, the knee joint of rats was detected by X-ray, the width of the knee joint was measured, oxidative stress indexes and inflammatory factors levels in serum and joint lavage fluid were detected, the morphology of the knee joint was observed by hematoxylin-eosin staining, full-thickness cartilage and hyaline cartilage thickness were measured, and the content of articular cartilage proteoglycan was observed by safranin O-fast green staining. The "drug-component-disease-target" network was constructed. Gene ontology enrichment analysis and Kyoto encyclopedia of genes and genomes enrichment analysis of effective targets were performed, and protein-protein interaction network maps were constructed using cytoscape software. RESULTS AND CONCLUSION: Sambucus adnata Wall extract could reduce tumor necrosis factor-α level in joint lavage fluid and serum levels of prostaglandin E2 and malondialdehyde, while increase the level of superoxide dismutase; relieve joint swelling caused by osteoarthritis; improve the histopathological state of articular cartilage, maintain the thickness of full-thickness articular cartilage, hyaline cartilage and the area of bone trabeculae in subchondral bone; and effectively prevent the loss of proteoglycans in articular cartilage and have a chondroprotective effect. Network pharmacological results showed that the methanol extract of Sambucus adnata Wall may have some targets related to inhibition of tumor necrosis factor, AKT1, interleukin-6, MAPK3, and SRC as well as inhibition of over-activation of EGFR signaling pathway and AGE-RAGE signaling pathway. [ABSTRACT FROM AUTHOR]

Knee osteoarthritis (KOA) is a chronic joint disease that significantly affects the health of the elderly. As an herbal remedy, Gubi decoction (GBD) has been traditionally used for the treatment of osteoarthritis‐related syndromes. However, the anti‐KOA efficacy and mechanism of GBD remain unclear. This study aimed to experimentally investigate the anti‐KOA efficacy and the underlying mechanism of GBD. The medial meniscus (DMM) mice model and IL‐1β‐stimulated chondrocytes were, respectively, constructed as in vivo and in vitro models of KOA to evaluate the osteoprotective effect and molecular mechanism of GBD. The UPLC–MS/MS analysis showed that GBD mainly contained pinoresinol diglucoside, rehmannioside D, hesperidin, liquiritin, baohuoside I, glycyrrhizic acid, kaempferol and tangeretin. Animal experiment showed that GBD could alleviate articular cartilage destruction and recover histopathological alterations in DMM mice. In addition, GBD inhibited chondrocyte apoptosis and restored DMM‐induced dysregulated autophagy evidenced by the upregulation of ATG7 and LC3 II/LC3 I but decreased P62 level. Mechanistically, METTL3‐mediated m6A modification decreased the expression of ATG7 in DMM mice, as it could be significantly attenuated by GBD. METTL3 overexpression significantly counteracted the protective effect of GBD on chondrocyte autophagy. Further research showed that GBD promoted proteasome‐mediated ubiquitination degradation of METLL3. Our findings suggest that GBD could act as a protective agent against KOA. The protective effect of GBD may result from its promotion on chondrocyte autophagy by suppressing METTL3‐dependent ATG7 m6A methylation. [ABSTRACT FROM AUTHOR]

Background: Osteoarthritis (OA) is a degenerative joint disease characterized by the progressive degeneration of articular cartilage, leading to pain, stiffness, and loss of joint function. The pathogenesis of OA involves multiple factors, including increased intracellular reactive oxygen species (ROS), enhanced chondrocyte apoptosis, and disturbances in cartilage matrix metabolism. These processes contribute to the breakdown of the extracellular matrix (ECM) and the loss of cartilage integrity, ultimately resulting in joint damage and dysfunction. RNA interference (RNAi) therapy has emerged as a promising approach for the treatment of various diseases, including hATTR and acute hepatic porphyria. By harnessing the natural cellular machinery for gene silencing, RNAi allows for the specific inhibition of target genes involved in disease pathogenesis. In the context of OA, targeting key molecules such as matrix metalloproteinase-13 (MMP13), which plays a critical role in cartilage degradation, holds great therapeutic potential. Results: In this study, we developed an innovative therapeutic approach for OA using a combination of liposome-encapsulated siMMP13 and NG-Monomethyl-L-arginine Acetate (L-NMMA) to form an injectable hydrogel. The hydrogel served as a delivery vehicle for the siMMP13, allowing for sustained release and targeted delivery to the affected joint. Experiments conducted on destabilization of the medial meniscus (DMM) model mice demonstrated the therapeutic efficacy of this composite hydrogel. Treatment with the hydrogel significantly inhibited the degradation of cartilage matrix, as evidenced by histological analysis showing preserved cartilage structure and reduced loss of proteoglycans. Moreover, the hydrogel effectively suppressed intracellular ROS accumulation in chondrocytes, indicating its anti-oxidative properties. Furthermore, it attenuated chondrocyte apoptosis, as demonstrated by decreased levels of apoptotic markers. Conclusion: In summary, the injectable hydrogel containing siMMP13, endowed with anti-ROS and anti-apoptotic properties, may represent an effective therapeutic strategy for osteoarthritis in the future. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a common joint disease and has been studied extensively in recent years as no promising therapy available so far for its treatment and remains a great challenge for health care specialists. Although the identification of some major mechanisms that contribute to this disease suggests a plethora of bioactive agents in tackling the associated complications yet OA's pathophysiology is still poorly understood owing to complex mechanistic changes observed. Experimental research is now exploring a wide range of therapeutically effective agents in an effort to find a way to repair OA-related joint degeneration and halt it from getting worse. Data was acquired and reviewed from most relevant and recent studies. This review summarizes the studies that are currently available and focuses on how various unconventional functional oils affect osteoarthritis and the affected joint tissues. An analysis of the recent scientific literature allowed us to highlight the potential anti-arthritic properties of edible oils and their main constituents, which seems to suggest an interesting new potential therapeutic application. Due to eccentric nature of OA, it is necessary to concentrate initially on the management of symptoms. The evidence supporting functional oils chondroprotective potential is still accumulating, underpinning a global need for more sustainable natural sources of treatment. More clinical research that focuses on the consequences of long-term treatment, possible negative effects, and epigenetic implications is necessary to get optimistic results. However, different animal or clinical studies suggest that linolenic and linoleic fatty acids decreased chondrocyte oxidative stress, cartilage breakdown, and expression of inflammatory markers. Distinct fatty acids along with minor components of oils also reduced the generation of prostaglandins and decreased oxidative stress. Furthermore, the potential roles of the main components of edible oils and possible negative results (if any) are also reported. While no severe side effects have been reported for any edible oils. Overall, these studies identify and support the use of functional oils as an adjuvant therapy for the management of OA and as a means of symptomatic alleviation for OA patients. However, to prove the effectiveness or to draw precise conclusions, high-quality clinical trials are required. [ABSTRACT FROM AUTHOR]

Skeletal dysplasias are group of rare genetic diseases resulting from mutations in genes encoding structural proteins of the cartilage extracellular matrix (ECM), signaling molecules, transcription factors, epigenetic modifiers, and several intracellular proteins. Cell division, organelle maintenance, and intracellular transport are all orchestrated by the cytoskeleton-associated proteins, and intracellular processes affected through microtubule-associated movement are important for the function of skeletal cells. Among microtubule-associated motor proteins, kinesins in particular have been shown to play a key role in cell cycle dynamics, including chromosome segregation, mitotic spindle formation, and ciliogenesis, in addition to cargo trafficking, receptor recycling, and endocytosis. Recent studies highlight the fundamental role of kinesins in embryonic development and morphogenesis and have shown that mutations in kinesin genes lead to several skeletal dysplasias. However, many questions concerning the specific functions of kinesins and their adaptor molecules as well as specific molecular mechanisms in which the kinesin proteins are involved during skeletal development remain unanswered. Here we present a review of the skeletal dysplasias resulting from defects in kinesins and discuss the involvement of kinesin proteins in the molecular mechanisms that are active during skeletal development. [ABSTRACT FROM AUTHOR]

[Display omitted] • Using genetic mouse models and a synthetic small molecule activator to demonstrate that Piezo1 activation promotes OA development and progression. • Utilizing biochemical and histological approaches to reveal that Piezo1 activation promotes Runx2 expression and chondrocyte catabolism through a PI3K-AKT dependent manner. • Demonstrated that artemisinin protects against OA lesions by inactivating Piezo1 in mice. Osteoarthritis (OA) is a devastating whole-joint disease affecting a large population worldwide with no cure; its mechanism remains poorly defined. Abnormal mechanical stress is the main pathological factor of OA. To investigate the effects of Piezo1 activation on OA development and progression and to explore Piezo1-targeting OA treatment. The expression levels of Piezo1 were determined in human OA cartilage and experimental OA mice. Mice with genetic Piezo1 deletion in chondrocytes or intra-articular injection of the Piezo1 activator Yoda1 were utilized to determine the effects on DMM-induced OA progression. Effects of artemisinin (ART), a potent antimalarial drug, on Piezo1 activation, chondrocyte metabolism and OA lesions were determined. Piezo1 expression was elevated in articular chondrocytes in human OA and DMM-induced mouse OA cartilage. Piezo1 deletion in chondrocytes largely attenuates DMM-induced OA-like phenotypes. In contrast, intra-articular injection of Yoda1 aggravates the knee joint OA lesions in mice. PIEZO1 activation increases, while PIEZO1 siRNA knockdown decreases, expression of RUNX2 and catabolic enzymes MMP13 and ADAMTS5 in primary human articular chondrocytes in a PI3K-AKT dependent manner. We have provided strong evidence supporting that ART is a novel and potent inhibitor of Piezo1 activation in primary OA-HACs and all cell lines examined, including human endothelial HUVEC cells, ATDC5 chondrocyte-like cells and MLO-Y4 osteocytes-like cells. Results from in vitro experiments confirmed that ART decreases the Yoda1-induced increases in the levels of OA-related genes and p-PI3K and p-AKT proteins in OA-HACs and alleviates DMM-induced OA lesions in mice. We establish a critical role of Piezo1 in promoting OA development and progression and define ART as a potential OA treatment. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a widespread inflammatory joint disease with significant global disability burden. Cuproptosis, a newly identified mode of cell death, has emerged as a crucial factor in various pathological conditions, including OA. In this context, our study aims to investigate the intrinsic relationship between cuproptosis‐related genes (CRGs) and OA, and assess their potential as biomarkers for OA diagnosis and treatment. Datasets from the GEO databases were analysed the differential expression of CRGs, leading to the identification of 10 key CRGs (CDKN2A, DLD, FDX1, GLS, LIAS, LIPT1, MTF1, PDHA1, DLAT and PDHB). A logistic regression analysis and calibration curves were used to show excellent diagnostic accuracy. Consensus clustering revealed two CRG patterns, with Cluster 1 indicating a closer association with OA progression. RT‐PCR confirmed a significant increase in the expression levels of these nine key genes in IL‐1β‐induced C28/i2 cells, and the expression of CDKN2A and FDX1 were also elevated in conditioned monocytes, while the expression of GLS and MTF1 were significantly decreased. In vitro experiments demonstrated that the expression levels of these 7/10 CRGs were significantly increased in chondrocytes induced by IL‐1β, and upon stimulation with cuproptosis inducers, chondrocyte apoptosis was exacerbated, accompanied by an increase in the expression of cuproptosis‐related proteins. These further substantiated our research findings and indicated that the nine selected cuproptosis genes have high potential for application in the diagnosis of OA. [ABSTRACT FROM AUTHOR]

Abstract Objective This study investigated the protective effects of low fluid shear stress (FSS ≤ 2 dyn/cm²) against interleukin-1β (IL-1β)-induced chondrocyte apoptosis and explored the underlying molecular mechanisms. Methods Chondrocytes were cultured under four conditions: control, IL-1β stimulation, low FSS, and combined low FSS + IL-1β stimulation. Apoptosis was assessed using Hoechst staining and flow cytometry. Western blotting determined the expression of caspase-3 (CASP3), caspase-8 (CASP8), and NF-κB p65. Quantitative real-time PCR measured miR-143-3p expression. The roles of miR-143-3p and the extracellular signal-regulated kinase 5 (ERK5)/Krüppel-like factor 4 (KLF4) signaling pathway were further investigated using miR-143-3p mimics and inhibitors, an ERK5 inhibitor, and a KLF4 overexpression vector. Results IL-1β induced significant chondrocyte apoptosis, which was markedly inhibited by low FSS. Mechanistically, low FSS suppressed miR-143-3p expression, thereby enhancing ERK5 signaling. This activated ERK5 subsequently upregulated KLF4 expression, further mitigating IL-1β-induced damage. Importantly, miR-143-3p overexpression under low FSS conditions exacerbated IL-1β-induced apoptosis, while miR-143-3p inhibition attenuated it. Consistent with this, ERK5 inhibition augmented IL-1β-induced apoptosis, whereas KLF4 overexpression suppressed it. Conclusion Low FSS protects chondrocytes from IL-1β-induced apoptosis by suppressing miR-143-3p and activating the ERK5/KLF4 signaling pathway. This study reveals a novel mechanism by which mechanical stimulation protects cartilage.

In articular cartilage, zone-specific cellular morphology is a typical characteristic of cartilage tissue, which is related with chondrocyte function, inflammation and osteoarthritis (OA). Chondrocyte hypertrophic phenotype is a criticle physiological process which indicates a hallmark of chondrocyte terminal differentiation and bone formation. Thus, developing a in vitro cell culture system for dynamic regulation of single chondrocyte volume at a three-dimensional (3D) level is particularly necessary for understanding how physical cues of matrix microenvironment regulate chondrocyte fate and the degeneration of articular cartilage. Here, based on the soft lithography techniques, we have constructed well-defined single-cell 3D dynamic volume control system to recapitulate the physiological matrix microenvironment of single chondrocyte niche. The results of finite element analysis indicated that the stress and strain distribution in the cell culture region is homogeneous during the stretching process. Additionally, 3D dynamic volume expansion and compression of single cells in physiological or hyperphysiological can be realized in this cell culture system. Our device for single-cell 3D dynamic culture provides a microphysiological culture system for chondrocytes to explore the mechanisms of cartilage hypertrophy, as well as develops a new paradigm for functional cartilage tissue engineering and regenerative medicine.

Abstract Low back pain (LBP) is largely attributed to intervertebral disc degeneration (IVDD), of which the endplate changes are an important component. However, the alterations in cell fate and properties within the endplates during degeneration remain unknown. Here, we firstly performed the single-cell RNA-sequencing analysis (scRNA-seq) of the cells focusing on degenerative human endplates. By unsupervised clustering of the 8,534 single-cell based on the gene expression, we identified nine distinct cell types. We employed Gene ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analysis, and the single-cell regulatory network inference and clustering (SCENIC) to determine the enriched pathways and transcriptional activities across seven chondrocyte subpopulations. Furthermore, two cell fates of chondrocyte differentiation were found by trajectory analysis, one was enriched in inflammation-related genes, and the other was related to extracellular matrix (ECM). Additionally, the intercellular interactions of macrophages (MA) and chondrocytes, T cells/natural killer cells (T/NK) and chondrocytes were examined by ligand-receptor pairs analysis, showing the important regulative function of FN1 from MA and CD74 from T/NK during endplate degeneration. Overall, our findings provide novel perspectives on the endplate degeneration at the single-cell level and a whole-transcriptome size.

Abstract Articular cartilage damage and wear can result in cartilage degeneration, ultimately culminating in osteoarthritis. Current surgical interventions offer limited capacity for cartilage tissue regeneration and offer only temporary alleviation of symptoms. Tissue engineering strategies are increasingly recognized as promising modalities for cartilage restoration. Currently, various biological scaffolds utilizing tissue engineering materials are extensively employed in both fundamental and clinical investigations of cartilage repair. In order to optimize the cartilage repair ability of tissue engineering scaffolds, researchers not only optimize the structure and properties of scaffolds from the perspective of materials science and manufacturing technology to enhance their histocompatibility, but also adopt strategies such as loading cells, cytokines, and drugs to promote cartilage formation. This review provides an overview of contemporary tissue engineering strategies employed in cartilage repair, as well as a synthesis of existing preclinical and clinical research. Furthermore, the obstacles faced in the translation of tissue engineering strategies to clinical practice are discussed, offering valuable guidance for researchers seeking to address these challenges.

Abstract Insulin-like growth factor (IGF) signaling is required for proper growth and skeletal development in vertebrates. Consequently, its dysregulation may lead to abnormalities of growth or skeletal structures. IGF is involved in the regulation of cell proliferation and differentiation of chondrocytes. However, the availability of bioactive IGF may be controlled by antagonizing IGF binding proteins (IGFBPs) in the circulation and tissues. As the metalloproteinase PAPP-A specifically cleaves members of the IGFBP family, we hypothesized that PAPP-A activity liberates bioactive IGF in cartilage. In PAPP-A knockout mice, the femur length was reduced and the mice showed a disorganized columnar organization of growth plate chondrocytes. Similarly, zebrafish lacking pappaa showed reduced length of Meckel’s cartilage and disorganized chondrocytes, reminiscent of the mouse knockout phenotype. Expression of chondrocyte differentiation markers (sox9a, ihha, and col10a1) was markedly affected in Meckel’s cartilage of pappaa knockout zebrafish, indicating that differentiation of chondrocytes was compromised. Additionally, the zebrafish pappaa knockout phenotype was mimicked by pharmacological inhibition of IGF signaling, and it could be rescued by treatment with exogenous recombinant IGF-I. In conclusion, our data suggests that IGF activity in the growing cartilage, and hence IGF signaling in chondrocytes, requires the presence of PAPP-A. The absence of PAPP-A causes aberrant chondrocyte organization and compromised growth in both mice and zebrafish.

Introduction: Osteoarthritis (OA) is a devastating whole-joint disease affecting a large population worldwide with no cure; its mechanism remains poorly defined. Abnormal mechanical stress is the main pathological factor of OA. Objectives: To investigate the effects of Piezo1 activation on OA development and progression and to explore Piezo1-targeting OA treatment. Methods: The expression levels of Piezo1 were determined in human OA cartilage and experimental OA mice. Mice with genetic Piezo1 deletion in chondrocytes or intra-articular injection of the Piezo1 activator Yoda1 were utilized to determine the effects on DMM-induced OA progression. Effects of artemisinin (ART), a potent antimalarial drug, on Piezo1 activation, chondrocyte metabolism and OA lesions were determined. Results: Piezo1 expression was elevated in articular chondrocytes in human OA and DMM-induced mouse OA cartilage. Piezo1 deletion in chondrocytes largely attenuates DMM-induced OA-like phenotypes. In contrast, intra-articular injection of Yoda1 aggravates the knee joint OA lesions in mice. PIEZO1 activation increases, while PIEZO1 siRNA knockdown decreases, expression of RUNX2 and catabolic enzymes MMP13 and ADAMTS5 in primary human articular chondrocytes in a PI3K-AKT dependent manner. We have provided strong evidence supporting that ART is a novel and potent inhibitor of Piezo1 activation in primary OA-HACs and all cell lines examined, including human endothelial HUVEC cells, ATDC5 chondrocyte-like cells and MLO-Y4 osteocytes-like cells. Results from in vitro experiments confirmed that ART decreases the Yoda1-induced increases in the levels of OA-related genes and p-PI3K and p-AKT proteins in OA-HACs and alleviates DMM-induced OA lesions in mice. Conclusions: We establish a critical role of Piezo1 in promoting OA development and progression and define ART as a potential OA treatment.

Osteoarthritis (OA) is the most prevalent degenerative joint disease worldwide. Effective management for early-stage OA is crucial. Denosumab (DS) has been widely used to treat osteoporosis (OP) and rheumatoid arthritis, but its potential for managing OA remains clear. We assessed the effects of DS on osteoclast activity and chondrocyte apoptosis using tartrate-resistant acid phosphatase (TRAP) assay, quantitative real-time polymerase chain reaction (qRT-PCR), flow cytometry, and TUNEL staining. To assess the impact of DS on the NF-κB pathway, we performed Western blot and immunofluorescence staining. Additionally, we used an OA model to explore the influence of DS on subchondral bone remodeling and cartilage degeneration in vivo. We found that DS hindered receptor activator of nuclear factor kappa B ligand (RANKL)-induced osteoclastogenesis by inhibiting the activity of the NF-κB pathway. Besides, DS alleviated reactive oxygen species (ROS)-induced apoptosis in chondrocytes by regulating the expression of genes related to apoptosis. Moreover, we observed an attenuation of OA-related subchondral bone remodeling and cartilage degeneration in vivo. Our findings indicate that DS could effectively suppress osteoclast activity and chondrocyte apoptosis, thereby mitigating OA-related subchondral bone remodeling and cartilage degeneration. These results provide a mechanistic basis for using DS to treat OA.

BACKGROUND: Currently, there is no drug that can completely cure osteoarthritis and its pathogenesis is still unclear. Circular RNAs (circRNAs) are differentially expressed in patients with osteoarthritis and are closely associated with various pathological processes in osteoarthritis. circRNAs play an important role in various physiological and pathological processes, such as chondrocyte homeostasis, extracellular matrix formation, and inflammatory response. OBJECTIVE: To mainly review the effects of circRNAs on pathological factors related to osteoarthritis, as well as the types and expression levels of circRNAs in osteoarthritis. METHODS: Related articles published from 1976 to August 2023 were retrieved from CNKI, WanFang, VIP, PubMed, Medline, Web of Science and Elsevier databases. The keywords were “osteoarthritis, circular RNA, non-coding RNA, synovial tissue, chondrocytes” in Chinese and English, respectively. All the relevant articles were screened, summarized, analyzed, and finally 69 papers were included in the review. RESULTS AND CONCLUSION: circRNAs are non-coding RNAs widely found in eukaryotic cells, with covalently closed continuous loop structure, but with no 5′ hat structure and 3′poly A tail, which are involved in multi-gene and multi-target regulatory networks and cannot be degraded by nucleic acid exonucleases (RNase R). circRNAs have a high abundance, high conservativeness and stability, and cell and tissue specificity. circRNAs have biological functions such as acting as molecular sponges for miRNAs, regulating linear RNA transcription and RNA shearing, interacting with RNA-bound proteins, and translating proteins. circRNAs regulate chondrocyte apoptosis and proliferation, degradation of cartilage extracellular matrix, and inflammation and other physiopathologic processes. circRNAs are expected to become biomarkers and potential therapeutic targets for clinical diagnosis and prognosis of osteoarthritis, and may become a new strategy for clinical treatment of osteoarthritis in the future. [ABSTRACT FROM AUTHOR]

BACKGROUND: Apoptosis is involved in the formation of degenerative joint diseases. Therefore, anti-chondrocyte apoptosis may be an effective way to treat osteoarthritis. Neferine has a wide range of pharmacological activities including anti-inflammatory, anti-tumor and anti-apoptotic activities, and its effect on OBJECTIVE: To investigate the effect of neferine on chondrocyte apoptosis induced by interleukin-1β and elucidate its possible mechanism. METHODS: (1) The rat chondrocytes at logarithmical stage were taken and intervened in five groups. The control group was cultured routinely. The model group was routinely cultured for 24 hours after treatment with interleukin-1β for 2 hours. The low-, medium-, and high-dose groups were treated with interleukin-1β for 2 hours and then cultured with 5, 10, and 20 μmol/L neferine for 24 hours, respectively. At the end of culture, cell apoptosis, chondroglycoprotein 39 and type II collagen levels in cell supernatants, mRNA and protein expression of apoptosis-related proteins, and mRNA and protein expression of proteins related to the protein kinase R-like endoplasmic reticulum kinase (PERK)/activating transcription factor 4 (ATF4) signaling pathway were detected. (2) Rat chondrocytes at logarithmic growth period were taken and divided into four groups: control group and model group were treated with the same intervention as above, drug group was treated with interleukin-1β for 2 hours and then cultured with 20 μmol/L neferine for 24 hours, and activator group was treated with interleukin1β for 2 hours and then cultured with 20 μmol/L neferine and CCT020312, an activator of PERK/ATF4 signaling pathway, for 24 hours. At the end of culture, cell proliferation was detected by cell counting kit-8 assay, apoptosis was detected by flow cytometry, and mRNA and protein expressions of apoptosis-related proteins and PERK/ATF4 signaling pathway-related proteins were detected. RESULTS AND CONCLUSION: (1) Compared with the control group, the model group showed increased apoptosis (P < 0.05), decreased proliferative activity (P < 0.05), increased level of chondroglycoprotein 39 (P < 0.05), decreased level of type II collagen (P < 0.05), decreased mRNA and protein expression of Bcl-2 protein (P < 0.05), increased mRNA and protein expression of Bax protein, increased caspase-3 mRNA expression, increased Cleaved-caspase-3 protein expression (P < 0.05), increased mRNA expression of PERK, ATF4, and C/EBP homologous protein (P < 0.05), and increased protein expression of p-PERK, ATF4, and C/EBP homologous protein (P < 0.05). Compared with the model group, neferine reversed the above effects of interleukin-1β on chondrocytes in a concentration-dependent manner. (2) Compared with the drug group, the activator group showed increased apoptosis (P < 0.05), decreased proliferative activity (P < 0.05), elevated mRNA expression of caspase-3, ATF4, and C/EBP homologous protein (P < 0.05), and elevated protein expression of Cleavedcaspase-3, ATF4, and C/EBP homologous protein (P < 0.05). (3) To conclude, neferine inhibits interleukin-1β-induced chondrocyte apoptosis and enhances cell proliferation activity, and the mechanism of action may be related to blocking the PERK/ATF4 signaling pathway in the endoplasmic reticulum. [ABSTRACT FROM AUTHOR]

BACKGROUND: Acupotomy is an effective method for the clinical treatment of osteoarthritis, with affirmed clinical outcomes, but the specific mechanisms remain unclear OBJECTIVE: To investigate the role of acupotomy in modulating chondrocyte autophagy to promote chondrocyte homeostasis in osteoarthritis. METHODS: Twenty-eight New Zealand rabbits were randomly divided into control group, osteoarthritis group, acupotomy group, and hyaluronic acid group, with seven rabbits in each group. The knee osteoarthritis rabbit model was prepared using the Videman method in the latter three groups. After modeling, the control group and osteoarthritis group received no interventions. The acupotomy group received acupotomy treatment 15 minutes per time, once a week, while the hyaluronic acid group received intra-articular injection of hyaluronic acid once a week, with a continuous treatment duration of 5 weeks. The day after the final intervention, knee joint macrostructure was observed using DR imaging, chondrocyte ultrastructure was examined through transmission electron microscopy, apoptosis of chondrocytes was assessed using Tunel staining, and western blot analysis was used to detect the expression of proteins related to the PI3K/Akt/mTOR pathway. RESULTS AND CONCLUSION: The DR imaging results revealed that the osteoarthritis group exhibited narrowed knee joint spaces and the formation of periarticular osteophytes, while the hyaluronic acid group and acupotomy group showed widened knee joint spaces with a reduction in periarticular osteophytes. Transmission electron microscopy results demonstrated a decreased number of autophagosomes in chondrocytes in the osteoarthritis group, along with nuclear shrinkage, nuclear membrane rupture, incomplete organelle morphology, and a clear tendency towards cell death. In contrast, both the hyaluronic acid group and acupotomy group exhibited a significant increase in autophagosomes, intact nuclear membranes, and a well-preserved cellular state. Tunel staining results indicated a considerable decrease in the number of apoptotic cells in the hyaluronic acid group and acupotomy group compared with the osteoarthritis group. Western blot results revealed that, compared with the control group, the expression levels of Beclin1, Cath D, and LC3II/LC3I were significantly decreased in the osteoarthritis group (P < 0.05), while the expression levels of p-Akt/Akt and p-mTOR/mTOR were significantly increased ( P < 0.05); compared with the osteoarthritis group, the expression levels of Beclin1, Cath D, and LC3II/LC3I were significantly increased in both the hyaluronic acid group and acupotomy group (P < 0.05), while the expression levels of p-Akt/Akt and p-mTOR/mTOR were significantly decreased (P < 0.05). To conclude, acupotomy intervention can modulate the PI3K/Akt/mTOR signaling pathway to enhance the autophagic level in chondrocytes, thereby maintaining chondrocyte homeostasis. This ultimately leads to a slowdown in cartilage degeneration. [ABSTRACT FROM AUTHOR]

BACKGROUND: Laser therapy is a non-invasive and painless treatment that is considered to be an effective method suitable for the treatment of osteoarthritis due to its simplicity and non-invasive nature. Currently, the mechanism of action of laser therapy is unclear and the results of studies on its clinical application are controversial. OBJECTIVE: To review and summarize the latest research progress of laser therapy on chondrocytes, animal experiments and clinical efficacy, and to explore the possible mechanism of laser-mediated multi-pathway biological effects, so as to provide a theoretical basis for further research on the laser treatment of osteoarthritis of the knee joint. METHODS: A literature search was performed in CNKI, WanFang Data, VIP and PubMed databases for relevant literature published from 2018 to 2023, with “laser therapy, low level laser therapy, high level laser therapy, photobiomodulation, knee osteoarthritis, chondrocytes” as the search terms in Chinese and English, respectively. Together with 14 articles searched manually, 70 articles were finally included for review. RESULTS AND CONCLUSION: Laser therapy in the treatment of knee osteoarthritis is mainly categorized into two types: low-level laser therapy and highlevel laser therapy. Differences in laser parameters and treatment protocols have a direct impact on laser efficacy. When appropriate parameters are used, low-level lasers show positive effects in cellular experiments, animal models, and clinical efficacy. High-level lasers have been less studied in the treatment of knee osteoarthritis, but some preliminary clinical studies have shown positive results. Cell experiments have shown that low-level laser promotes chondrocyte proliferation and cartilage matrix synthesis, thereby reducing inflammatory response. Animal experiments have shown that low-level laser can reduce the release of pro-inflammatory factors, promote cartilage matrix synthesis, inhibit matrix degradation, and effectively improve the repair process of cartilage tissue. Low-level laser is also able to reduce oxidative stress damage and relieve pain in knee osteoarthritis. In clinical trials, both low- and high-level laser can reduce patients’ pain and improve functional activities. The combination of laser therapy and exercise therapy modalities may improve the therapeutic effect. Lasers may affect intracellular signaling and cellular functions through photobiological or thermodynamic effects. This provides direct evidence that laser promotes articular cartilage regeneration. [ABSTRACT FROM AUTHOR]

BACKGROUND: The results of in vivo and in vitro studies showed that catalpol from Rehmannia glutinosa can significantly reduce the level of inflammatory indexes in the synovial tissue of rats with knee osteoarthritis, and meanwhile, it can delay the progression of knee osteoarthritis. But whether catalpol from Rehmannia glutinosa affects chondrocyte senescence and then delay the progression of knee osteoarthritis has not yet been clarified. OBJECTIVE: To investigate investigate whether catalpol from Rehmannia glutinosa could regulate ATDC5 chondrocyte senescence and the possible mechanisms. METHODS: ATDC5 chondrocytes were divided into blank group (0.1% bovine serum albumin), model group (0.1% bovine serum albumin+1 µmol/L adriamycin), low-dose catalpol group (0.1% bovine serum albumin+1 µmol/L adriamycin+20 µmol/L catalpol from Rehmannia glutinosa) and high-dose catalpol group (0.1% bovine serum albumin+1 µmol/L adriamycin+80 µmol/L catalpol from Rehmannia glutinosa). Adriamycin-induced ATDC5 chondrocyte senescence model was constructed, and the corresponding treatments were given according to the above groups. Cell counting kit-8 assay was used to detect the effects of catalpol from Rehmannia glutinosa on ATDC5 chondrocyte viability, and to screen the optimal concentration of catalpol from Rehmannia glutinosa. The senescence of ATDC5 chondrocytes in each group was detected by β-galactosidase staining after the corresponding treatments. Real-time fluorescence quantitative PCR and western blot were used to detect the mRNA and protein expression of P21, P53, type II collagen, matrix metalloproteinase 13, and interleukin-6. Immunofluorescence method was used to detect the expression of P21, P53 and type II collagen. Flow cytometry was used to detect apoptosis in each group. RESULTS AND CONCLUSION: ATDC5 chondrocytes were identified to be successfully induced and senescence model was induced. Catalpol from Rehmannia glutinosa at the concentrations of 0, 20, 40, and 80 µmol/L showed no significant effects on the cell viability, suggesting that catalpol from Rehmannia glutinosa is non-cytotoxic and can be used safely (P > 0.05); when the concentration was ≥ 100 µmol/L, the cell viability was reduced, suggesting that there may be cytotoxic. Therefore, 80 µmol/L was chosen as the high dose for subsequent experiments in this study. The percentage of positive cells in the model group was (86.93±2.18)%, which was significantly higher than that in the blank group [(17.32±0.72)%; P < 0.05]. Compared with the model group, the percentage of positive cells was significantly lower in the low- and high-dose catalpol groups [(57.28±1.73)% and (27.18±0.97)%, respectively; both P < 0.05]. Compared with the model group, the relative expression of P21, P53, matrix metalloproteinase 13, and interleukin-6 at mRNA and protein levels was significantly downregulated in the low- and high-dose catalpol groups, while the relative expression of type II collagen at mRNA and protein levels was significantly upregulated in both groups (P < 0.05), especially in the high-dose catalpol group (P < 0.05). Compared with the model group, the fluorescence intensities of P21 and P53 were significantly weakened in the low- and high-dose catalpol groups, while the fluorescence intensity of type II collagen was significantly enhanced in the low- and high-dose catalpol groups (P < 0.05), especially in the high-dose catalpol group (P < 0.05). The cell apoptosis detected by Annexin V/PI method showed that there was no significant difference between the model group and the blank group (P > 0.05); compared with the model group, the apoptotic index was significantly elevated in the low- and high-dose catalpol groups, especially in the high-dose catalpol group (P < 0.05). To conclude, catalpol from Rehmannia glutinosa can slow the progression of osteoarthritis by promoting apoptosis of senescent ATDC5 chondrocytes, further removing senescent ATDC5 chondrocytes, and decreasing the senescence-associated phenotypes. [ABSTRACT FROM AUTHOR]

BACKGROUND: The content of serum thrombin in patients with osteoarthritis is significantly higher than that in normal individuals, and thrombin can induce inflammatory degeneration of rat chondrocytes, suggesting that inhibiting the function of thrombin may become a method for treating osteoarthritis. Celecoxib is a common therapeutic drug for the clinical treatment of osteoarthritis. It is not yet known whether it improves chondrocyte degeneration by inhibiting the activity of thrombin. OBJECTIVE: To investigate the effect of celecoxib on thrombin-induced degeneration of rat chondrocytes. METHODS: Thrombin levels in the serum of osteoarthritis patients and normal individuals were detected by an ELISA kit. Primary chondrocytes of neonatal Sprague-Dawley rats were isolated, and all experiments were performed with cells from passage one. Chondrocytes were randomly divided into three groups: control group, thrombin group, and celecoxib group. The cell morphology of the three groups was observed under an inverted microscope, and an Edu kit was used to detect the cell proliferation. qRT-PCR was used to detect the expression of extracellular matrix components (aggrecan, elastin, cartilage oligomeric matrix proteins), inflammatory factors (interleukin-1, interleukin-6, and tumor necrosis factor-α), and chemokines (monocyte chemotactic protein 2, monocyte chemotactic protein 7, granulocyte chemotactic protein 6). The expression of type 2 collagen α1 was detected by immunofluorescence. Western blot method was used to detect the expression of catabolic metabolism genes, such as matrix metalloproteinase 9, matrix metalloproteinase 13, and cyclooxygenase 2. RESULTS AND CONCLUSION: Patients with osteoarthritis had higher levels of thrombin in the serum compared with normal individuals. Under the microscope, celecoxib was found to significantly inhibit fibroid changes in chondrocytes. Compared with the thrombin group, celecoxib inhibited the proliferation of chondrocytes. The downregulation of extracellular matrix gene expression, such as type II collagen α1, in the thrombin group was inhibited by celecoxib (P < 0.05). Thrombin promoted the expression of inflammatory factors (interleukin-1, interleukin-6, and tumor necrosis factor-α), chemokines (monocyte chemotactic protein 2, monocyte chemotactic protein 7, granulocyte chemotactic protein 6), as well as catabolic genes (matrix metalloproteinase 9, matrix metalloproteinase 13, and cyclooxygenase 2), and under the intervention of celecoxib, the expression of these genes could be downregulated (P < 0.05). Overall, these findings indicate that celecoxib inhibits the pro-inflammatory effects of thrombin and thereby ameliorates chondrocyte degeneration in rats. [ABSTRACT FROM AUTHOR]

Age-related deterioration of condylar cartilage is an etiological factor in temporomandibular joint-osteoarthritis (TMJ-OA). However, its underlying mechanism remains unknown. Therefore, we examined age-related changes and the relationship between mTOR signaling and primary cilia in condylar cartilage to determine the intrinsic mechanisms of age-related TMJ-OA. Age-related morphological changes were analyzed using micro-computed tomography and safranin O-stained histological samples of the mandibular condyle of C57BL/6J mice (up to 78 weeks old). Immunohistochemistry was used to assess the activity of mTOR signaling, primary cilia frequency, and Golgi size of condylar chondrocytes. Four-week-old mice receiving an 11-week series of intraperitoneal injections of rapamycin, a potent mTOR signaling inhibitor, were used for the histological evaluation of the condylar cartilage. The condylar cartilage demonstrated an age-related reduction in cartilage area, including chondrocyte size, cell density, and cell size distribution. The Golgi size, primary cilia frequency, and mTOR signaling also decreased with age. Rapamycin injections resulted in both diminished cartilage area and cell size, resembling the phenotypes observed in aged mice. Rapamycin-injected mice also exhibited a smaller Golgi size and lower primary cilia frequency in condylar cartilage. We demonstrated that a loss of primary cilia due to a decline in mTOR signaling was correlated with age-related deteriorations in condylar cartilage. Our findings provide new insights into the tissue homeostasis of condylar cartilage, contributing to understanding the etiology of age-related TMJ-OA. [ABSTRACT FROM AUTHOR]

Inflammation is a complex biological process protecting the body from diverse external threats. Effectively performing this task requires an intricate, well‐regulated interplay of different cells and tissues. Furthermore, several cells participating in inflammation can assume diverse phenotypes. A classic and relatively well‐studied example of phenotypic diversity in inflammation is macrophage polarization. Based on the TH1/TH2 phenotypes of T helper cells, this scheme has proinflammatory "classical/M1" activation contrasted with the anti‐inflammatory and healing‐promoting "alternative/M2" phenotype. Some authors have extended the concept into an M17 phenotype induced by the classic TH17 cytokine IL‐17. Phenotypic changes in chondrocytes have also been studied especially in the context of osteoarthritis (OA), and there are indications that these cells can also assume polarized phenotypes at least partly analogous to those of TH cells and macrophages. The therapeutic success of biological agents targeting TH1/TH2/TH17 inductor and/or effector cytokines displays the utility of the concept of polarization. The aim of this focused review is to survey the internal and external factors affecting macrophage and chondrocyte phenotypes (such as inflammatory cytokines, widely used medications and natural products) and to explore the possibility of ameliorating pathological states by modulating these phenotypes. [ABSTRACT FROM AUTHOR]

Mammalian somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) via the forced expression of Yamanaka reprogramming factors. However, only a limited population of the cells that pass through a particular pathway can metamorphose into iPSCs, while the others do not. This study aimed to clarify the pathways that chondrocytes follow during the reprogramming process. The fate of human articular chondrocytes under reprogramming was investigated through a time-coursed single-cell transcriptomic analysis, which we termed an inverse genetic approach. The iPS interference technique was also employed to verify that chondrocytes inversely return to pluripotency following the proper differentiation pathway. We confirmed that human chondrocytes could be converted into cells with an iPSC phenotype. Moreover, it was clarified that a limited population that underwent the silencing of SOX9 , a master gene for chondrogenesis, at a specific point during the proper transcriptome transition pathway, could eventually become iPSCs. Interestingly, the other cells, which failed to be reprogrammed, followed a distinct pathway toward cells with a surface zone chondrocyte phenotype. The critical involvement of cellular communication network factors (CCNs) in this process was indicated. The idea that chondrocytes, when reprogrammed into iPSCs, follow the differentiation pathway backward was supported by the successful iPS interference using SOX9. This inverse genetic strategy may be useful for seeking candidates for the master genes for the differentiation of various somatic cells. The utility of CCNs in articular cartilage regeneration is also supported. [ABSTRACT FROM AUTHOR]