Mume Fructus reduces interleukin-1 beta-induced cartilage degradation via MAPK downregulation in rat articular chondrocytes.

Osteoarthritis is the most prevalent type of degenerative arthritis. It is characterized by persistent pain, joint dysfunction, and physical disability. Pain relief and inflammation control are prioritised during osteoarthritis treatment Mume Fructus (Omae), a fumigated product of the Prunus mume fruit, is used as a traditional medicine in several Asian countries. However, its therapeutic mechanism of action and effects on osteoarthritis and articular chondrocytes remain unknown. In this study, we analyzed the anti-osteoarthritis and articular regenerative effects of Mume Fructus extract on rat chondrocytes. Mume Fructus treatment reduced the interleukin-1ß-induced expression of matrix metalloproteinase 3, matrix metalloproteinase 13, and a disintegrin and metalloproteinase with thrombospondin type 1 motifs 5. Additionally, it enhanced collagen type II alpha 1 chain and aggrecan accumulation in rat chondrocytes. Furthermore, Mume Fructus treatment regulated the inflammatory cytokine levels, mitogen-activated protein kinase phosphorylation, and nuclear factor-kappa B activation. Overall, our results demonstrated that Mume Fructus inhibits osteoarthritis progression by inhibiting the nuclear factor-kappa B and mitogen-activated protein kinase pathways to reduce the levels of inflammatory cytokines and prevent cartilage degeneration. Therefore, Mume Fructus may be a potential therapeutic option for osteoarthritis.

Chondroitin Sulfate/Hyaluronic Acid-Blended Hydrogels Suppress Chondrocyte Inflammation under Pro-Inflammatory Conditions.

Osteoarthritis is characterized by enzymatic breakdown of the articular cartilage via the disruption of chondrocyte homeostasis, ultimately resulting in the destruction of the articular surface. Decades of research have highlighted the importance of inflammation in osteoarthritis progression, with inflammatory cytokines shifting resident chondrocytes into a pro-catabolic state. Inflammation can result in poor outcomes for cells implanted for cartilage regeneration. Therefore, a method to promote the growth of new cartilage and protect the implanted cells from the pro-inflammatory cytokines found in the joint space is required. In this study, we fabricate two gel types: polymer network hydrogels composed of chondroitin sulfate and hyaluronic acid, glycosaminoglycans (GAGs) known for their anti-inflammatory and prochondrogenic activity, and interpenetrating networks of GAGs and collagen I. Compared to a collagen-only hydrogel, which does not provide an anti-inflammatory stimulus, chondrocytes in GAG hydrogels result in reduced production of pro-inflammatory cytokines and enzymes as well as preservation of collagen II and aggrecan expression. Overall, GAG-based hydrogels have the potential to promote cartilage regeneration under pro-inflammatory conditions. Further, the data have implications for the use of GAGs to generally support tissue engineering in pro-inflammatory environments.

FTH1 protects against osteoarthritis by MAPK pathway inhibition of extracellular matrix degradation.

OBJECTIVE: Ferritin heavy chain 1 (FTH1) is an important subunit of ferro-storing proteins and is indispensable for iron metabolism. Though it has been extensively studied in numerous organs and diseases, the relationship between FTH1 and osteoarthritis (OA) is unclear. DESIGN: Primary murine chondrocytes and cartilage explants were treated with FTH1 siRNA for 72 h. Mice were injected with adenovirus expressing FTH1 after destabilized medial meniscus (DMM) surgery. These approaches were used to determine the effect of FTH1 expression on the pathophysiology of OA. RESULTS: FTH1 expression was down regulated in OA patients and mice after DMM surgery. Knock down of FTH1 induced articular cartilage damage and extracellular matrix degradation in cartilage explants. Further, over expression of FTH1 reduced the susceptibility of chondrocytes to ferroptosis and reversed decrements in SOX9 and aggrecan after DMM surgery. Moreover, FTH1 relieved OA by inhibition of the chondrocyte MAPK pathway. CONCLUSION: This study found FTH1 to play an essential role in extracellular matrix degradation, ferroptosis, and chondrocytes senescence during OA progression. Further, injection of adenovirus expressing FTH1 may be a potential strategy for OA prevention and therapy.

Genotype and phenotype in patients with ACAN gene variants: Three cases and literature review.

OBJECTIVE: To characterize the phenotype spectrum, diagnosis, and response to growth-promoting therapy in patients with ACAN variants causing familial short stature. METHODS: Three families with ACAN variants causing short stature were reported. Similar cases in the literature were summarized, and the genotype and phenotype were analyzed. RESULTS: Three novel heterozygous variants, c.757+1G>A, (splicing), c.6229delG, p.(Asp2078Tfs*1), and c.6679C>T, p.(Gln2227*) in the ACAN gene were identified. A total of 314 individuals with heterozygous variants from 105 families and 8 individuals with homozygous variants from 4 families were confirmed to have ACAN variants from literature and our 3 cases. Including our 3 cases, the variants reported comprised 33 frameshift, 39 missense, 23 nonsense, 5 splicing, 4 deletion, and 1 translocation variants. Variation points are scattered throughout the gene, while exons 12, 15, and 10 were most common (25/105, 11/105, and 10/105, respectively). Some identical variants existing in different families could be hot variants, c.532A>T, p.(Asn178Tyr), c.1411C>T, p.(Gln471*), c.1608C>A, p.(Tyr536*), c.2026+1G>A, (splicing), and c.7276G>T, p.(Glu2426*). Short stature, early-onset osteoarthritis, brachydactyly, midfacial hypoplasia, and early growth cessation were the common phenotypic features. The 48 children who received rhGH (and GnRHa) treatment had a significant height improvement compared with before (-2.18 ± 1.06 SD vs. -2.69 ± 0.95 SD, p < 0.001). The heights of children who received rhGH (and GnRHa) treatment were significantly improved compared with those of untreated adults (-2.20 ± 1.10 SD vs. -3.24 ± 1.14 SD, p < 0.001). CONCLUSION: Our study achieves a new understanding of the phenotypic spectrum, diagnosis, and management of individuals with ACAN variants. No clear genotype-phenotype relationship of patients with ACAN variants was found. Gene sequencing is necessary to diagnose ACAN variants that cause short stature. In general, appropriate rhGH and/or GnRHa therapy can improve the adult height of affected pediatric patients caused by ACAN variants.

Deficiency of Cbfß in articular cartilage leads to osteoarthritis-like phenotype through Hippo/Yap, TGFß, and Wnt/ß-catenin signaling pathways.

Osteoarthritis (OA) is the most prevalent degenerative joint disorder, causing physical impairments among the elderly. Core binding factor subunit ß (Cbfß) has a critical role in bone homeostasis and cartilage development. However, the function and mechanism of Cbfß in articular cartilage and OA remains unclear. We found that Cbfßf/fAggrecan-CreERT mice with Cbfß-deficiency in articular cartilage developed a spontaneous osteoarthritis-like phenotype with articular cartilage degradation. Immunofluorescence staining showed that Cbfßf/fAggrecan-CreERT mice exhibited a significant increase in the expression of articular cartilage degradation markers and inflammatory markers in the knee joints. RNA-sequencing analysis demonstrated that Cbfß orchestrated Hippo/Yap, TGFß/Smad, and Wnt/ß-catenin signaling pathways in articular cartilage, and Cbfß deficiency resulted in the abnormal expression of downstream genes involved in maintaining articular cartilage homeostasis. Immunofluorescence staining results showed Cbfß deficiency significantly increased active ß-catenin and TCF4 expression while reducing Yap, TGFß1, and p-Smad 2/3 expression. Western blot and qPCR validated gene expression changes in hip articular cartilage of Cbfß-deficient mice. Our results demonstrate that deficiency of Cbfß in articular cartilage leads to an OA-like phenotype via affecting Hippo/Yap, TGFß, and Wnt/ß-catenin signaling pathways, disrupting articular cartilage homeostasis and leading to the pathological process of OA in mice. Our results indicate that targeting Cbfß may be a potential therapeutic target for the design of novel and effective treatments for OA.

Role of Galectin-3 in intervertebral disc degeneration: an experimental study.

BACKGROUND: This study investigated the role of Galectin-3 in the degeneration of intervertebral disc cartilage. METHODS: The patients who underwent lumbar spine surgery due to degenerative disc disease were recruited and divided into Modic I, Modic II, and Modic III; groups. HE staining was used to detect the pathological changes in endplates. The changes of Galectin-3, MMP3, Aggrecan, CCL3, and Col II were detected by immunohistochemistry, RT-PCR, and Western blot. MTT and flow cytometry were used to detect cartilage endplate cell proliferation, cell cycle, and apoptosis. RESULTS: With the progression of degeneration (from Modic I to III), the chondrocytes and density of the cartilage endplate of the intervertebral disc decreased, and the collagen arrangement of the cartilage endplate of the intervertebral disc was broken and calcified. Meanwhile, the expressions of Aggrecan, Col II, Galectin-3, Aggrecan, and CCL3 gradually decreased. After treatment with Galectin-3 inhibitor GB1107, the proliferation of rat cartilage end plate cells was significantly reduced (P < 0.05). GB1107 (25 µmol/L) also significantly promoted the apoptosis of cartilage endplate cells (P < 0.05). Moreover, the percentage of cartilage endplate cells in the G1 phase was significantly higher, while that in the G2 and S phases was significantly lower (P < 0.05). Additionally, the mRNA and protein expression levels of MMP3, CCL3, and Aggrecan in rat cartilage end plate cells were lower than those in the control group. CONCLUSIONS: Galectin-3 decreases with the progression of the cartilage endplate degeneration of the intervertebral disc. Galectin-3 may affect intervertebral disc degeneration by regulating the degradation of the extracellular matrix.

Whole-Transcriptome Sequencing of Knee Joint Cartilage from Kashin-Beck Disease and Osteoarthritis Patients.

The aim of this study was to provide a comprehensive understanding of similarities and differences in mRNAs, lncRNAs, and circRNAs within cartilage for Kashin-Beck disease (KBD) compared to osteoarthritis (OA). We conducted a comparison of the expression profiles of mRNAs, lncRNAs, and circRNAs via whole-transcriptome sequencing in eight KBD and ten OA individuals. To facilitate functional annotation-enriched analysis for differentially expressed (DE) genes, DE lncRNAs, and DE circRNAs, we employed bioinformatic analysis utilizing Gene Ontology (GO) and KEGG. Additionally, using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), we validated the expression levels of four cartilage-related genes in chondrocytes. We identified a total of 43 DE mRNAs, 1451 DE lncRNAs, and 305 DE circRNAs in KBD cartilage tissue compared to OA (q value < 0.05; |log2FC| > 1). We also performed competing endogenous RNA network analysis, which identified a total of 65 lncRNA-mRNA interactions and 4714 miRNA-circRNA interactions. In particular, we observed that circRNA12218 had binding sites for three miRNAs targeting ACAN, while circRNA12487 had binding sites for seven miRNAs targeting COL2A1. Our results add a novel set of genes and non-coding RNAs that could potentially serve as candidate diagnostic biomarkers or therapeutic targets for KBD patients.

Development of alginate-collagen interpenetrating network for osteoarthritic cartilage by in situ softening.

This study presents an alginate-collagen interpenetrating network (IPN) matrix of incorporating collagen fibrils into an alginate hydrogel by physical mixing and controlled gelation. The resulting matrix closely mimics the physiological and pathological stiffness range of the chondrocyte pericellular matrix (PCM). Chondrocytes were cultured within three-dimensional (3D) alginate-collagen IPN matrices with varying stiffness, namely Firm, Medium, and Soft. Alginate lyase was introduced to study the effects of the changes in stiffness of the Firm on chondrocyte response by in situ softening. The developed alginate-collagen IPN matrix displayed good cell-biocompatibility. Compared with stiffer tissue culture plastic (TCP), chondrocytes grown within Firm displayed a stabilized differentiated phenotype characterized by higher expression levels of aggrecan, collagen II, and SOX-9. Moreover, the developed alginate-collagen IPN matrix exhibited a gradually increased percentage of propidium iodide (PI)-positive dead cells with decreasing stiffness. Softer matrices directed cells towards higher proliferation rates and spherical morphologies while stimulating chondrocyte cluster formation. Furthermore, reducing Firm stiffness by in situ softening decreased aggrecan expression, contributing to matrix degradation similar to that seen in osteoarthritis (OA). Hence, the 3D alginate-collagen IPN constructs hold significant potential for in vitro replicating PCM stiffness changes observed in OA cartilage.

Evaluating the Anti-Osteoarthritis Potential of Standardized Boswellia serrata Gum Resin Extract in Alleviating Knee Joint Pathology and Inflammation in Osteoarthritis-Induced Models.

Osteoarthritis is a widespread chronic degenerative disease marked by the deterioration of articular cartilage, modifications in subchondral bone, and a spectrum of symptoms, including pain, stiffness, and disability. Ultimately, this condition impairs the patient's quality of life. This study aimed to evaluate the therapeutic efficacy of standardized Boswellia serrata gum resin extract (BSRE) in a rat model of monosodium iodoacetate (MIA)-induced osteoarthritis. A total of 60 rats were allocated into six groups: normal control group (NC), osteoarthritis control (injected with MIA, OC), O + B50 (injected with MIA and treated with 50 mg/kg body weight (BW) BSRE), O + B75 (injected with MIA and treated with 75 mg/kg BW BSRE), O + B100 (injected with MIA and treated with 100 mg/kg BW BSRE), and O + M (injected with MIA and treated with 150 mg/kg BW methyl sulfonyl methane). Several parameters, including knee joint swelling, histopathological changes, and the expression of collagen type II alpha 1 (COL2A1) and aggrecan, were comprehensively assessed. Concurrently, the serum levels and mRNA expression of inflammatory mediators, cytokines, and matrix metalloproteinases (MMPs) were analyzed in both the serum and knee joint synovium. The results demonstrated that BSRE significantly mitigated knee joint swelling, cartilage destruction, and tissue deformation. Notably, BSRE administration markedly upregulated the expression of COL2A1 and aggrecan while concurrently reducing levels of nitric oxide, prostaglandin E2, leukotriene B4, interleukin (IL)-6, and tumor necrosis factor (TNF)-α. Furthermore, a substantial decrease was observed in the mRNA expression of inducible nitric oxide synthase, cyclooxygenase-2, 5-lipoxygenase, IL-6, TNF-α and MMP-3 and -13, thereby indicating promising therapeutic implications for osteoarthritis. In conclusion, BSRE exhibited anti-inflammatory properties and inhibited cartilage matrix degradation in a rat model of MIA-induced osteoarthritis, with the O + B100 group showing significant reductions in swelling and notable improvements in joint cartilage damage. These findings illuminate the preventive and therapeutic potential of BSRE for osteoarthritis treatment, emphasizing the criticality of exhaustive evaluation of novel compounds.

Ginsenoside Rg1 relieves rat intervertebral disc degeneration and inhibits IL-1ß-induced nucleus pulposus cell apoptosis and inflammation via NF-κB signaling pathway.

The study aimed to investigate the effect of ginsenoside Rg1 on intervertebral disc degeneration (IVDD) in rats and IL-1ß-induced nucleus pulposus (NP) cells, and explore its underlying mechanism. Forty IVDD rat models were divided into the IVDD group, low-dose (L-Rg1) group (intraperitoneal injection of 20 mg/kg/d ginsenoside Rg1), medium-dose (M-Rg1) group (intraperitoneal injection of 40 mg/kg/d ginsenoside Rg1), and high-dose (H-Rg1) group (intraperitoneal injection of 80 mg/kg/d ginsenoside Rg1). The pathological change was observed by HE and safranin O-fast green staining. The expression of IL-1ß, IL-6, TNF-α, MMP3, aggrecan, and collagen II was detected. The expression of NF-κB p65 in IVD tissues was detected. Rat NP cells were induced by IL-1ß to simulate IVDD environment and divided into the control group, IL-1ß group, and 20, 50, and 100 µmol/L Rg1 groups. The cell proliferation activity, the apoptosis, and the expression of IL-6, TNF-α, MMP3, aggrecan, collagen II, and NF-κB pathway-related protein were detected. In IVDD rats, ginsenoside Rg1 improved the pathology of IVD tissues; suppressed the expression of IL-1ß, IL-6, TNF-α, aggrecan, and collagen II; and inhibited the expression of p-p65/p65 and nuclear translocation of p65, to alleviate the IVDD progression. In the IL-1ß-induced NP cells, ginsenoside Rg1 also improved the cell proliferation and inhibited the apoptosis and the expression of IL-6, TNF-α, aggrecan, collagen II, p-p65/p65, and IκK in a dose-dependent manner. Ginsenoside Rg1 alleviated IVDD in rats and inhibited apoptosis, inflammatory response, and ECM degradation in IL-1ß-induced NP cells. And Rg1 may exert its effect via inhibiting the activation of NF-κB signaling pathway.

Inhibition of cc chemokine receptor 10 ameliorates osteoarthritis via inhibition of the phosphoinositide-3-kinase/Akt/mammalian target of rapamycin pathway.

BACKGROUND: Osteoarthritis (OA) is a joint disease characterized by inflammation and progressive cartilage degradation. Chondrocyte apoptosis is the most common pathological feature of OA. Interleukin-1ß (IL-1ß), a major inflammatory cytokine that promotes cartilage degradation in OA, often stimulates primary human chondrocytes in vitro to establish an in vitro OA model. Moreover, IL-1ß is involved in OA pathogenesis by stimulating the phosphoinositide-3-kinase (PI3K)/Akt and mitogen-activated protein kinases pathways. The G-protein-coupled receptor, cc chemokine receptor 10 (CCR10), plays a vital role in the occurrence and development of various malignant tumors. However, the mechanism underlying the role of CCR10 in the pathogenesis of OA remains unclear. We aimed to evaluate the protective effect of CCR10 on IL-1ß-stimulated CHON-001 cells and elucidate the underlying mechanism. METHODS: The CHON-001 cells were transfected with a control small interfering RNA (siRNA) or CCR10-siRNA for 24 h, and stimulated with 10 ng/mL IL-1ß for 12 h to construct an OA model in vitro. The levels of CCR10, cleaved-caspase-3, MMP-3, MMP-13, Collagen II, Aggrecan, p-PI3K, PI3K, p-Akt, Akt, phosphorylated-mammalian target of rapamycin (p-mTOR), and mTOR were detected using quantitative reverse transcription polymerase chain reaction and western blotting. Viability, cytotoxicity, and apoptosis of CHON-001 cells were assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, lactate dehydrogenase assay (LDH), and flow cytometry analysis, respectively. Inflammatory cytokines (TNF-α, IL-6, and IL-8) were assessed using enzyme-linked immunosorbent assay. RESULTS: Level of CCR10 was substantially higher in the IL-1ß-stimulated CHON-001 cells than that in the control group, whereas CCR10 was down-regulated in the CCR10-siRNA transfected CHON-001 cells compared to that in the control-siRNA group. Notably, CCR10 inhibition alleviated IL-1ß-induced inflammatory injury in the CHON-001 cells, as verified by enhanced cell viability, inhibited LDH release, reduced apoptotic cells, and cleaved-caspase-3 expression. Meanwhile, IL-1ß induced the release of tumor necrosis factor alpha, IL-6, and IL-8, increase of MMP-3 and MMP-13, and decrease of Collagen II and Aggrecan in the CHON-001 cells, which were reversed by CCR10-siRNA. However, these effects were reversed upon PI3K agonist 740Y-P treatment. Further, IL-1ß-induced PI3K/Akt/mTOR signaling pathway activation was inhibited by CCR10-siRNA, which was increased by 740Y-P treatment. CONCLUSION: Inhibition of CCR10 alleviates IL-1ß-induced chondrocytes injury via PI3K/Akt/mTOR pathway inhibition, suggesting that CCR10 might be a promising target for novel OA therapeutic strategies.

Menstrual blood-derived mesenchymal stem cells combined with collagen I gel as a regenerative therapeutic strategy for degenerated disc after discectomy in rats.

BACKGROUND: Annulus fibrosis (AF) defects have been identified as the primary cause of disc herniation relapse and subsequent disc degeneration following discectomy. Stem cell-based tissue engineering offers a promising approach for structural repair. Menstrual blood-derived mesenchymal stem cells (MenSCs), a type of adult stem cell, have gained attention as an appealing source for clinical applications due to their potential for structure regeneration, with ease of acquisition and regardless of ethical issues. METHODS: The differential potential of MenSCs cocultured with AF cells was examined by the expression of collagen I, SCX, and CD146 using immunofluorescence. Western blot and ELISA were used to examine the expression of TGF-ß and IGF-I in coculture system. An AF defect animal model was established in tail disc of Sprague-Dawley rats (males, 8 weeks old). An injectable gel containing MenSCs (about 1*106/ml) was fabricated and transplanted into the AF defects immediately after the animal model establishment, to evaluate its repairment properties. Disc degeneration was assessed via magnetic resonance (MR) imaging and histological staining. Immunohistochemical analysis was performed to assess the expression of aggrecan, MMP13, TGF-ß and IGF-I in discs with different treatments. Apoptosis in the discs was evaluated using TUNEL, caspase3, and caspase 8 immunofluorescence staining. RESULTS: Coculturing MenSCs with AF cells demonstrated ability to express collagen I and biomarkers of AF cells. Moreover, the coculture system presented upregulation of the growth factors TGF-ß and IGF-I. After 12 weeks, discs treated with MenSCs gel exhibited significantly lower Pffirrmann scores (2.29 ± 0.18), compared to discs treated with MenSCs (3.43 ± 0.37, p < 0.05) or gel (3.71 ± 0.29, p < 0.01) alone. There is significant higher MR index in disc treated with MenSCs gel than that treated with MenSCs (0.51 ± 0.05 vs. 0.24 ± 0.04, p < 0.01) or gel (0.51 ± 0.05 vs. 0.26 ± 0.06, p < 0.01) alone. Additionally, MenSCs gel demonstrated preservation of the structure of degenerated discs, as indicated by histological scoring (5.43 ± 0.43 vs. 9.71 ± 1.04 in MenSCs group and 10.86 ± 0.63 in gel group, both p < 0.01), increased aggrecan expression, and decreased MMP13 expression in vivo. Furthermore, the percentage of TUNEL and caspase 3-positive cells in the disc treated with MenSCs Gel was significantly lower than those treated with gel alone and MenSCs alone. The expression of TGF-ß and IGF-I was higher in discs treated with MenSCs gel or MenSCs alone than in those treated with gel alone. CONCLUSION: MenSCs embedded in collagen I gel has the potential to preserve the disc structure and prevent disc degeneration after discectomy, which was probably attributed to the paracrine of growth factors of MenSCs.

Composite Zonal Scaffolds of Collagen I/II for Meniscus Regeneration.

The meniscus is divided into three zones according to its vascularity: an external vascularized red-red zone mainly comprising collagen I, a red-white interphase zone mainly comprising collagens I and II, and an internal white-white zone rich in collagen II. Known scaffolds used to treat meniscal injuries do not reflect the chemical composition of the vascular areas of the meniscus. Therefore, in this study, four composite zonal scaffolds (named A, B, C, and D) were developed and characterized; the developed scaffolds exhibited the main chemical components of the external (collagen I), interphase (collagens I/II), and internal (collagen II) zones of the meniscus. Noncomposite scaffolds were also produced (named E), which had the same shape as the composite scaffolds but were entirely made of collagen I. The composite zonal scaffolds were prepared using different concentrations of collagen I and the same concentration of collagen II and were either cross-linked with genipin or not cross-linked. Porous, biodegradable, and hydrophilic scaffolds with an expected chemical composition were obtained. Their pore size was smaller than the size reported for the meniscus substitutes; however, all scaffolds allowed the adhesion and proliferation of human adipose-derived stem cells (hADSCs) and were not cytotoxic. Data from enzymatic degradation and hADSC proliferation assays were considered for choosing the cross-linked composite scaffolds along with the collagen I scaffold and to test if composite zonal scaffolds seeded with hADSC and cultured with differentiation medium produced fibrocartilage-like tissue different from that formed in noncomposite scaffolds. After 21 days of culture, hADSCs seeded on composite scaffolds afforded an extracellular matrix with aggrecan, whereas hADSCs seeded on noncomposite collagen I scaffolds formed a matrix-like fibrocartilage without aggrecan.

TMF inhibits extracellular matrix degradation by regulating the C/EBPß/ADAMTS5 signaling pathway in osteoarthritis.

Osteoarthritis (OA) is a chronic joint disease, characterized by degenerative destruction of articular cartilage. Chondrocytes, the unique cell type in cartilage, mediate the metabolism of extracellular matrix (ECM), which is mainly constituted by aggrecan and type II collagen. A disintegrin and metalloproteinase with thrombospondin 5 (ADAMTS5) is an aggrecanase responsible for the degradation of aggrecan in OA cartilage. CCAAT/enhancer binding protein ß (C/EBPß), a transcription factor in the C/EBP family, has been reported to mediate the expression of ADAMTS5. Our previous study showed that 5,7,3',4'-tetramethoxyflavone (TMF) could activate the Sirt1/FOXO3a signaling in OA chondrocytes. However, whether TMF protected against ECM degradation by down-regulating C/EBPß expression was unknown. In this study, we found that aggrecan expression was down-regulated, and ADAMTS5 expression was up-regulated. Knockdown of C/EBPß could up-regulate aggrecan expression and down-regulate ADAMTS5 expression in IL-1ß-treated C28/I2 cells. TMF could compromise the effects of C/EBPß on OA chondrocytes by activating the Sirt1/FOXO3a signaling. Conclusively, TMF exhibited protective activity against ECM degradation by mediating the Sirt1/FOXO3a/C/EBPß pathway in OA chondrocytes.

Bone protective effect of sinomenine against monosodium iodoacetate induced knee and hip injury in rat model: an inflammatory pathway.

PURPOSE: Osteoarthritis (OA) is a degenerative joint disease which is categorized via destruction of joint cartilage and it also affects the various joints, especially knees and hips. Sinomenine active phytoconstituents isolated from the stem of Sinomenium acutum and already proof anti-inflammatory effect against the arthritis model of rodent. In this experimental protocol, we scrutinized the anti-osteoarthritis effect of sinomenine against monosodium iodoacetate (MIA) induced OA in rats. METHODS: MIA (3 mg/50 µL) was used for inducing the OA in the rats, and rats received the oral administration of sinomenine (2.5, 5 and 7.5 mg/kg body weight) up to the end of the experimental study (four weeks). The body and organs weight were estimated. Aggrecan, C-terminal cross-linked telopeptide of type II collagen (CTX-II), glycosaminoglycans (GCGs), monocyte chemoattractant protein-1 (MCP-1), Interferon gamma (IFN-γ), antioxidant, inflammatory cytokines, inflammatory mediators and matrix metalloproteinases (MMP) were analyzed. RESULTS: Sinomenine significantly (P < 0.001) boosted the body weight and reduced the heart weight, but the weight of spleen and kidney remain unchanged. Sinomenine significantly (P < 0.001) reduced the level of nitric oxide, MCP-1 and improved the level of aggrecan, IFN-γ and GCGs. Sinomenine remarkably upregulated the level of glutathione, superoxide dismutase and suppressed the level of malonaldehyde. It effectually modulated the level of inflammatory cytokines and inflammatory mediators and significantly (P < 0.001) reduced the level of MMPs, like MMP-1, 2, 3, 9 and 13. CONCLUSIONS: Sinomenine is a beneficial active agent for the treatment of OA disease.

Passaged Articular Chondrocytes From the Superficial Zone and Deep Zone Can Regain Zone-Specific Properties After Redifferentiation.

BACKGROUND: Bioengineered cartilage is a developing therapeutic to repair cartilage defects. The matrix must be rich in collagen type II and aggrecan and mechanically competent, withstanding compressive and shearing loads. Biomechanical properties in native articular cartilage depend on the zonal architecture consisting of 3 zones: superficial, middle, and deep. The superficial zone chondrocytes produce lubricating proteoglycan-4, whereas the deep zone chondrocytes produce collagen type X, which allows for integration into the subchondral bone. Zonal and chondrogenic expression is lost after cell number expansion. Current cell-based therapies have limited capacity to regenerate the zonal structure of native cartilage. HYPOTHESIS: Both passaged superficial and deep zone chondrocytes at high density can form bioengineered cartilage that is rich in collagen type II and aggrecan; however, only passaged superficial zone-derived chondrocytes will express superficial zone-specific proteoglycan-4, and only passaged deep zone-derived chondrocytes will express deep zone-specific collagen type X. STUDY DESIGN: Controlled laboratory study. METHODS: Superficial and deep zone chondrocytes were isolated from bovine joints, and zonal subpopulations were separately expanded in 2-dimensional culture. At passage 2, superficial and deep zone chondrocytes were seeded, separately, in scaffold-free 3-dimensional culture within agarose wells and cultured in redifferentiation media. RESULTS: Monolayer expansion resulted in loss of expression for proteoglycan-4 and collagen type X in passaged superficial and deep zone chondrocytes, respectively. By passage 2, superficial and deep zone chondrocytes had similar expression for dedifferentiated molecules collagen type I and tenascin C. Redifferentiation of both superficial and deep zone chondrocytes led to the expression of collagen type II and aggrecan in both passaged chondrocyte populations. However, only redifferentiated deep zone chondrocytes expressed collagen type X, and only redifferentiated superficial zone chondrocytes expressed and secreted proteoglycan-4. Additionally, redifferentiated deep zone chondrocytes produced a thicker and more robust tissue compared with superficial zone chondrocytes. CONCLUSION: The recapitulation of the primary phenotype from passaged zonal chondrocytes introduces a novel method of functional bioengineering of cartilage that resembles the zone-specific biological properties of native cartilage. CLINICAL RELEVANCE: The recapitulation of the primary phenotype in zonal chondrocytes could be a possible method to tailor bioengineered cartilage to have zone-specific expression.

Vitamin D delays intervertebral disc degeneration and improves bone quality in ovariectomized rats.

Known to be involved in bone-cartilage metabolism, Vitamin D (VD) may play a role in human's disc pathophysiology. Given that postmenopausal women are prone to suffer VD deficiency and intervertebral disc degeneration (IDD), this study is intended to investigate whether VD can delay IDD in ovariectomized rats by improving bone microstructure and antioxidant stress. Female Sprague-Dawley rats were randomly allocated into four groups: sham, oophorectomy (OVX)+VD deficiency (VDD), OVX, and OVX+VD supplementation (VDS). In vivo, after a 6-month intervention, imaging and pathology slice examinations showed that IDD induced by OVX was significantly alleviated in VDS and deteriorated by VDD. The expressions of aggrecan and Collagen II in intervertebral disc were reduced by OVX and VDD, and elevated by VDS. Compared with the OVX+VDD and OVX group vertebrae, OVX+VDS group vertebrae showed significantly improved endplate porosity and lumbar bone mineral density with increased percent bone volume and trabecular thickness. Furthermore, 1α,25(OH)2D3 restored the redox balance (total antioxidant capacity, ratio of oxidized glutathione/glutathione) in the disc. The cocultivation of 1α,25(OH)2D3 and nucleus pulposus cells (NPCs) was conducted to observe its potential ability to resist excessive oxidative stress damage induced by H2O2. In vitro experiments revealed that 1α,25(OH)2D3 reduced the senescence, apoptosis, and extracellular matrix degradation induced by H2O2 in NPCs. In conclusion, VDS exhibits protective effects in OVX-induced IDD, partly by regulating the redox balance and preserving the microstructure of endplate. This finding provides a new idea for the prevention and treatment of IDD.

Dose- and stage-dependent toxic effects of prenatal prednisone exposure on fetal articular cartilage development.

Prednisone is frequently used to treat rheumatoid diseases in pregnant women because of its high degree of safety. Whether prenatal prednisone exposure (PPE) negatively impacts fetal articular cartilage development is unclear. In this study, we simulated a clinical prednisone treatment regimen to examine the effects of different timings and doses of PPE on cartilage development in female and male fetal mice. Prednisone doses (0.25, 0.5, and 1 mg/kg/d) was administered to Kunming mice at different gestational stages (0-9 gestational days, GD0-9), mid-late gestation (GD10-18), or during the entire gestation (GD0-18) by oral gavage. The amount of matrix aggrecan (ACAN) and collagen type II a1(COL2a1), and expression of transforming growth factor ß1 (TGFß1) signaling pathway also demonstrated that the chondrocyte count and ACAN and COL2a1 expression reduced in fetal mice with early and mid-late PPE, with the reduction being more significant in the mice with early PPE than that in those with PPE at other stages. Prenatal exposure to different prednisone doses prevented the reduction of TGFß signaling pathway-related genes [TGFßR1, SMAD family member 3 (Smad3), SRY-box9 (SOX9)] as well as ACAN and COL2a1 mRNA expression levels in fetal mouse cartilage, with the most significant decrease after 1 mg/kg·d PPE. In conclusion, PPE can inhibit/restrain fetal cartilage development, with the greatest effect at higher clinical dose (1 mg/kg·d) and early stage of pregnancy (GD0-9), and the mechanism may be related to TGFß signaling pathway inhibition. The result of this study provide a theoretical and experimental foundation for the rational clinical use of prednisone.

Association of cartilage metabolism biomarkers and 25(OH)D levels with muscle biomechanical functions in professional rowers and canoeists.

The purpose of the study was to assess the association of cartilage metabolism biomarkers and vitamin D metabolite levels with muscle biomechanical functions in professional rowers and canoeists. The serum levels of aggrecan, cartilage oligomeric matrix protein (COMP), and 25-hydroxyvitamin D (25(OH)D) were determined in elite male sweep-oar rowers (n = 24) and canoeists (n = 15). This was followed by a biomechanical study consisting in isometric measurement of peak torque (PT) of muscles involved in the rowing cycle in the athletes. There were found significant correlations of COMP with the ratio of trunk PT flexor to extensor (p < 0.05) and 25(OH)D with trunk PT-left rotators (p < 0.05), knee joints PT-left and right flexor (p ≤ 0.01), ratio of knee joint PT-right flexor to knee joint PT-right extensor (p < 0.05) in rowers and aggreccan with elbow joint PT of the right flexor (p ≤ 0.01) and extensor (p = 0.05) in canoeists. The correlations of COMP and aggrecan levels with PT of the muscle groups studied in rowers and canoeists indicate the importance of stabilizing the muscular system in cartilage metabolism. The relationship between 25(OH)D status and biomechanical parameters confirm that vitamin D plays an important role in maintaining skeletal muscle health.

Microscale strain concentrations in tissue-engineered osteochondral implants are dictated by local compositional thresholds and architecture.

Tissue-engineered osteochondral implants manufactured from condensed mesenchymal stem cell bodies have shown promise for treating focal cartilage defects. Notably, such manufacturing techniques have shown to successfully recapture the bulk mechanical properties of native cartilage. However, the relationships among the architectural features, local composition, and micromechanical environment within tissue-engineered cartilage from cell-based aggregates remain unclear. Understanding such relationships is crucial for identifying critical parameters that can predict in vivo performance. Therefore, this study investigated the relationship among architectural features, composition, and micromechanical behavior of tissue-engineered osteochondral implants. We utilized fast-confocal microscopy combined with a strain mapping technique to analyze the micromechanical behavior under quasi-static loading, as well as Fourier Transform Infrared Spectroscopy to analyze the local compositions. More specifically, we investigated the architectural features and compositional distributions generated from tissue maturation, along with macro- and micro-level strain distributions. Our results showed that under compression, cell-based aggregates underwent deformation followed by body movement, generating high local strain around the boundaries, where local aggrecan concentration was low and local collagen concentration was high. By analyzing the micromechanics and composition at the single aggregate length scale, we identified a strong threshold relationship between local strain and compositions. Namely at the aggrecan concentration below 0.015 arbitrary unit (A.U.) and the collagen concentration above 0.15 A.U., the constructs experienced greater than threefold increase in compressive strain. Overall, this study suggests that local compositional features are the primary driver of the local mechanical environment in tissue-engineered cartilage constructs, providing insight into potential quality control parameters for manufacturing tissue-engineered constructs.