Mesenchymal stem cells (MSCs) are usually cultured in a normoxic atmosphere (21%) in vitro, while the oxygen concentrations in human tissues and organs are 1% to 10% when the cells are transplanted in vivo. However, the impact of hypoxia on MSCs has not been deeply studied, especially its translational application. In the present study, we investigated the characterizations of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) in hypoxic (1%) and normoxic (21%) atmospheres with a long-term culture from primary to 30 generations, respectively. The comparison between both atmospheres systematically analyzed the biological functions of MSCs, mainly including stemness maintenance, immune regulation, and resistance to chondrocyte apoptosis, and studied their joint function and anti-inflammatory effects in osteoarthritis (OA) rats constructed by collagenase II.Aims
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Aims. This study aimed to define the histopathology of degenerated humeral head cartilage and synovial inflammation of the glenohumeral joint in patients with omarthrosis (OmA) and cuff tear arthropathy (CTA). Additionally, the potential of immunohistochemical tissue biomarkers in reflecting the degeneration status of humeral head cartilage was evaluated. Methods. Specimens of the humeral head and synovial tissue from 12 patients with OmA, seven patients with CTA, and four body donors were processed histologically for examination using different histopathological scores. Osteochondral sections were immunohistochemically stained for collagen type I, collagen type II, collagen neoepitope C1,2C, collagen type X, and osteocalcin, prior to semiquantitative analysis. Matrix metalloproteinase (MMP)-1, MMP-3, and
Adenosine, lidocaine, and Mg2+ (ALM) therapy exerts differential immuno-inflammatory responses in males and females early after anterior cruciate ligament (ACL) reconstruction (ACLR). Our aim was to investigate sex-specific effects of ALM therapy on joint tissue repair and recovery 28 days after surgery. Male (n = 21) and female (n = 21) adult Sprague-Dawley rats were randomly divided into ALM or Saline control treatment groups. Three days after ACL rupture, animals underwent ACLR. An ALM or saline intravenous infusion was commenced prior to skin incision, and continued for one hour. An intra-articular bolus of ALM or saline was also administered prior to skin closure. Animals were monitored to 28 days, and joint function, pain, inflammatory markers, histopathology, and tissue repair markers were assessed.Aims
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Therapeutic agents that prevent chondrocyte loss, extracellular matrix (ECM) degradation, and osteoarthritis (OA) progression are required. The expression level of epidermal growth factor (EGF)-like repeats and discoidin I-like domains-containing protein 3 (EDIL3) in damaged human cartilage is significantly higher than in undamaged cartilage. However, the effect of EDIL3 on cartilage is still unknown. We used human cartilage plugs (ex vivo) and mice with spontaneous OA (in vivo) to explore whether EDIL3 has a chondroprotective effect by altering OA-related indicators.Aims
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Osteoarthritis (OA) is mainly caused by ageing, strain, trauma, and congenital joint abnormalities, resulting in articular cartilage degeneration. During the pathogenesis of OA, the changes in subchondral bone (SB) are not only secondary manifestations of OA, but also an active part of the disease, and are closely associated with the severity of OA. In different stages of OA, there were microstructural changes in SB. Osteocytes, osteoblasts, and osteoclasts in SB are important in the pathogenesis of OA. The signal transduction mechanism in SB is necessary to maintain the balance of a stable phenotype, extracellular matrix (ECM) synthesis, and bone remodelling between articular cartilage and SB. An imbalance in signal transduction can lead to reduced cartilage quality and SB thickening, which leads to the progression of OA. By understanding changes in SB in OA, researchers are exploring drugs that can regulate these changes, which will help to provide new ideas for the treatment of OA. Cite this article:
To explore the novel molecular mechanisms of histone deacetylase 4 (HDAC4) in chondrocytes via RNA sequencing (RNA-seq) analysis. Empty adenovirus (EP) and a Aims
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Osteoarthritis (OA) is a chronic degenerative joint disease characterized by progressive cartilage degradation, synovial membrane inflammation, osteophyte formation, and subchondral bone sclerosis. Pathological changes in cartilage and subchondral bone are the main processes in OA. In recent decades, many studies have demonstrated that activin-like kinase 3 (ALK3), a bone morphogenetic protein receptor, is essential for cartilage formation, osteogenesis, and postnatal skeletal development. Although the role of bone morphogenetic protein (BMP) signalling in articular cartilage and bone has been extensively studied, many new discoveries have been made in recent years around ALK3 targets in articular cartilage, subchondral bone, and the interaction between the two, broadening the original knowledge of the relationship between ALK3 and OA. In this review, we focus on the roles of ALK3 in OA, including cartilage and subchondral bone and related cells. It may be helpful to seek more efficient drugs or treatments for OA based on ALK3 signalling in future.
This study aimed to investigate the effect of irisin on human nucleus pulposus cells (hNPCs) in vitro. Our hypothesis was that irisin would improve hNPC metabolism and proliferation. hNPCs were isolated from intervertebral discs and cultured in alginate beads. hNPCs were exposed to phosphate-buffered saline (PBS) or recombinant irisin (r-irisin) at 5, 10 and 25 ng/mL (n=4). Each experiment was performed in triplicate. Cell proliferation was assessed with trypan blue staining-automated cell counting and PicoGreen assay. Glycosaminoglycan (GAG) content was measured using the DMMB assay. Metabolic activity was assessed with the MTT assay and the Griess Reagent System. Gene expression of collagen type II (COL2), matrix metalloproteinase (MMP)-13, tissue inhibitor of matrix metalloproteinase (TIMP)-1 and −3, aggrecan, interleukin (IL)-1β, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-5 was measured by RT-PCR. MTT assay and ADAMTS-5, COL2, TIMP-1 and IL-1β gene expression were evaluated following incubation with 5, 10 and 25 ng/mL r-irisin for 24 hours and subsequent culture with 10 ng/ml IL-1β and vice versa (incubation for 24 hours with IL-1β and subsequent culture with r-irisin). Irisin increased hNPC proliferation (p<0.001), metabolic activity (p<0.05), GAG content (p<0.01), as well as COL2 (p<0.01), aggrecan (p<0.05), TIMP-1 and −3 (p<0.01) gene expression, while decreasing
Regulation of articular cartilage homeostasis is a complex process in which biologic and mechanical factors are involved. Hyperactivation of Wnt signaling, associated with osteoarthritis (OA), could jeopardize the protective anabolic effect of physiological loading. Here, we investigated the role of excessive Wnt signalling in cartilage molecular responses to loading. Human cartilage explants were harvested from hips of donors without OA. The Wnt agonist CHIR99021 was used to activate Wnt signalling 24 hours before cartilage explants were subjected to a loading protocol consisting of 2 cycles of 1 hour of 10% compression at 1 Hz, followed by 1-hour free swelling. Mechano-responsiveness was evaluated using the expression of type II collagen, aggrecan and
Previous studies showed that telo-peptides degraded from type II collagen, a type of collagen fragments, could induce cartilage damage in bovine stifle joints. We aim to investigate the role of integrins (ITGs) and matrix metalloproteinases (MMPs) in collagen fragment-induced human cartilage damage that is usually observed in osteoarthritis (OA). We hypothesized that N-telopeptide (NT) derived from type II collagen could up-regulate the expression of β1 integrin (ITGB1) and then MMPs that may lead to osteoarthritic cartilage damage. Human chondrocytes were isolated from femoral head or tibial plateau of patients receiving arthroplasty (N = 24). Primary chondrocyte cultures were either treated with 30 µM NT, or 30 µM scrambled NT (SN), or PBS, or left untreated for 24 hrs. Total proteins and RNAs were extracted for examination of expression of ITGB1 and MMPs-3&13 with Western blotting and quantitative real-time PCR. Compared to untreated or PBS treated chondrocytes, NT-treated chondrocytes expressed significantly higher levels of ITGB1 and MMPs-3&-13. However, SN also up-regulated expression of ITGB1 and
TGF-β/Smad2 signaling is considered to be one of the important pathways involved in osteoarthritis (OA) and protein phosphatase magnesium-dependent 1A (PPM1A) functions as an exclusive phosphatase of Smad2 and regulates TGF-β signaling, here, we investigated the functional role of PPM1A in OA pathogenesis. PPM1A expressions in both human OA cartilage and experimental OA mice chondrocytes were analyzed immunohistochemically. Besides, the mRNA and protein expression of PPM1A induced by IL-1β treatment were also detected by q-PCR and immunofluorescence in vitro. OA was induced in PPM1A knockout (KO) mice by destabilization of the medial meniscus (DMM), and histopathological examination was performed. OA was also induced in wild-type (WT) mice, which were then treated with an intra-articular injection of a selective PPM1A inhibitor for 8 weeks. PPM1A protein expressions were increased in both human OA cartilage and experimental OA mice chondrocytes. We also found that treatment with IL-1β in mouse primary chondrocytes significantly increased both mRNA and protein expression of PPM1A in vitro. Importantly, our data showed that PPM1A deletion could substantially protect against surgically induced OA. Concretely, the average OARSI score and quantification of BV/TV of subchondral bone in KO mice were significantly lower than that in WT mice 8 weeks after DMM surgery. Besides, TUNEL staining revealed a significant decrease in apoptotic chondrocytes in PPM1A-KO mice with DMM operation. With OA induction, the rates of chondrocytes positive for
Osteoarthritis (OA) is a common age-related degenerative joint disease, affecting 7% of the global population, more than 500 million people worldwide. Exosomes from mesenchymal stem cells (MSCs) showed promise for OA treatment, but the insufficient biological targeting weakens its efficacy and might bring side effects. Here, we report the chondrocyte-targeted exosomes synthesized via click chemistry as a novel treatment for OA. Exosomes are isolated from human umbilical cord-derived MSCs (hUC-MSCs) using multistep ultracentrifugation process, and identified by electron microscope and nanoparticle tracking analysis (NTA). Chondrocyte affinity peptide (CAP) is conjugated on the surface of exosomes using click chemistry. For tracking, nontagged exosomes and CAP-exosomes are labeled by Dil, a fluorescent dye that highlights the lipid membrane of exosomes. To verify the effects of CAP-exosomes, nontagged exosomes and CAP-exosomes are added into the culture medium of interleukin (IL)-1β-induced chondrocytes. Immunofluorescence are used to test the expression of matrix metalloproteinase (MMP)-13. CAP-exosomes, compared with nontagged exosomes, are more easily absorbed by chondrocytes. What's more, CAP-exosomes induced lower
Rheumatoid arthritis (RA) is a common chronic immune disease. Berberine, as its main active ingredient, was also contained in a variety of medicinal plants such as Berberaceae, Buttercup, and Rutaceae, which are widely used in digestive system diseases in traditional Chinese medicine with anti-inflammatory and antibacterial effects. The aims of this article were to explore the therapeutic effect and mechanism of berberine on rheumatoid arthritis. Cell Counting Kit-8 was used to evaluate the effect of berberine on the proliferation of RA fibroblast-like synoviocyte (RA-FLS) cells. The effect of berberine on matrix metalloproteinase (MMP)-1, MMP-3, receptor activator of nuclear factor kappa-Β ligand (RANKL), tumour necrosis factor alpha (TNF-α), and other factors was determined by enzyme-linked immunoassay (ELISA) kit. Transcriptome technology was used to screen related pathways and the potential targets after berberine treatment, which were verified by reverse transcription-polymerase chain reaction (RT-qPCR) and Western blot (WB) technology.Aims
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The February 2023 Foot & Ankle Roundup360 looks at: Joint inflammatory response in ankle and pilon fractures; Tibiotalocalcaneal fusion with a custom cage; Topical application of tranexamic acid can reduce blood loss in calcaneal fractures; Risk factors for failure of total ankle arthroplasty; Pain catastrophizing: the same as pain forecasting?.
Circular RNA (circRNA) is involved in the regulation of articular cartilage degeneration induced by inflammatory factors or oxidative stress. In a previous study, we found that the expression of Minus RNA sequencing, fluorescence in situ hybridization, and quantitative real-time polymerase chain reaction (qRT-PCR) were used to detect the expression of Aims
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Intervertebral disc degeneration (IDD) affects more than 80% of the population all over the world. Current strategies for the treatment of IDD are based on conservative or surgical procedures with the aim of relieving pain. Mesenchymal stem cell (MSC) transplantation has emerged as a promising therapy in recent decades, but studies showed that the particularly hostile microenvironment in the intervertebral disc (IVD) can compromise cells survival rate. The use of exosomes, extracellular vesicles released by various cell types, possess considerable economic advantages including low immunogenicity and toxicity. Exosomes allow intercellular communication by conveying functional proteins, RNA, miRNA and lipids between cells. The purpose of this study is to assess the therapeutic effects of exosomes derived from Wharton Jelly mesenchymal stromal cells (WJ-MSC) on human nucleuspulposus cells (hNPC) in an in vitro 3D culture model. Exosomes (exos) were isolated by tangential flow filtration of WJ-MSC conditioned media and characterized by: quantification with BCA test; morphological observation with TEM, surface marker expression by WB and size evaluation by NTA. Confocal microscopy has been used to identify exosomes marked with PKH26 and monitor fusion and/or incorporation in hNPC. hNPC were isolated from waste surgical material from patients undergoing discectomy (n = 5), expanded, encapsulated in alginate beads and treated with: culture medium (control group); WJ-MSC exos (WJ-exos) at different concentrations (10 μg/ml, 50 μg/ml and 100 μg/ml). They were then analysed for: cell proliferation (Trypan Blu); viability (Live/Dead Assay); quantification of nitrites (Griess) and glycosaminoglycans, GAG (DMBB). The hNPC in alginate beads treated for 7 days were included in paraffin and histologically analysed to determine the presence of extracellular matrix (ECM) components. Finally, the expression levels of catabolic and anabolic genes were evaluated through real-time polymerase chain reaction (qPCR). All concentrations of WJ-exos under exam were capable to induce a significant increase in cell proliferation after 10 and 14 days of treatment (p < 0.01 and p < 0.001, respectively). Live/Dead assay showed a decrease in cell death at 50 μg/ml of WJ-exos (p < 0.05). While cellular oxidative stress indicator, nitrite production, was reduced in a dose-dependent way and statistically significant only with 100 μg/ml of WJ-exos (p < 0.05). WJ-exos at 10 and 100 μg/ml induced a significant increase in GAG content (p < 0.05; p < 0.01, respectively) confirmed by Alcian Blu staining. Exos derived from WJ-MSC modulated gene expression levels by increasing expression of ACAN and SOX-9 genes and reducing significantly of IL-6, MMP-1,
Introduction:. Exercise has showed to reduce pain and improve function in patients with discogenic low back pain (LBP). Although there is currently no biologic evidence that the intervertebral disc (IVD) can respond to physical exercise in humans, a recent study has shown that chronic running exercise is associated with increased IVD hydration and hypertrophy1. Irisin, a myokine released upon muscle contraction, has demonstrated to yield anabolic effects on different cell types, including chondrocytes2. This study aimed to investigate the effect of irisin on human nucleus pulposus cells (hNPCs). Our hypothesis is that irisin may improve hNPCs metabolism and proliferation. METHODS:. The hNPCs, isolated from discectomy surgical waste material (n = 5), were expanded and encapsulated in alginate beads. The hNPCs were treated with: i) only growth medium (control); ii) medium with recombinant irisin (r-IR) at different concentrations (5, 10 and 25 ng / mL); iii) medium with Interleukin-1β (IL1β); iv) medium with IL1β for 24 h and then with IL1β and r-IR; v) medium with r-IR for 24 h and then with r-IR and IL1 β. We evaluated proliferation (trypan blue and PicoGreen), metabolic activity (MTT), nitrite concentration (Griess), and expression levels of catabolic and anabolic genes via real-time polymerase chain reaction (qPCR). Each analysis was performed in triplicate for each donor and each experiment was performed three times. Data were expressed as mean ± S.D. One-way ANOVA was used for the groups under exam. RESULTS:. Irisin increased hNPCs proliferation (p < 0.001), metabolic activity at 10 ng/mL (p < 0.05), and GAG content at concentration of 10 ng/mL and 25 ng/mL (p < 0.01; p < 0.001, respectively). The production of nitrites, used as an indicator of cellular oxidative stress, was significantly decreased (p < 0.01). Gene expression levels compared to the control group increased for COL2A1 (p < 0.01), ACAN (p < 0.05), TIMP-1 and −3 (p < 0.01), while a decrease in mRNA levels of
Osteoarthritis (OA) is a common degenerative joint disease worldwide, which is characterized by articular cartilage lesions. With more understanding of the disease, OA is considered to be a disorder of the whole joint. However, molecular communication within and between tissues during the disease process is still unclear. In this study, we used transcriptome data to reveal crosstalk between different tissues in OA. We used four groups of transcription profiles acquired from the Gene Expression Omnibus database, including articular cartilage, meniscus, synovium, and subchondral bone, to screen differentially expressed genes during OA. Potential crosstalk between tissues was depicted by ligand-receptor pairs.Aims
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Autologous chondrocyte implantation (ACI) is a promising treatment for articular cartilage degeneration and injury; however, it requires a large number of human hyaline chondrocytes, which often undergo dedifferentiation during in vitro expansion. This study aimed to investigate the effect of suramin on chondrocyte differentiation and its underlying mechanism. Porcine chondrocytes were treated with vehicle or various doses of suramin. The expression of collagen, type II, alpha 1 (COL2A1), aggrecan (ACAN); COL1A1; COL10A1; SRY-box transcription factor 9 (SOX9); nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX); interleukin (IL)-1β; tumour necrosis factor alpha (TNFα); IL-8; and matrix metallopeptidase 13 (MMP-13) in chondrocytes at both messenger RNA (mRNA) and protein levels was determined by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and western blot. In addition, the supplementation of suramin to redifferentiation medium for the culture of expanded chondrocytes in 3D pellets was evaluated. Glycosaminoglycan (GAG) and collagen production were evaluated by biochemical analyses and immunofluorescence, as well as by immunohistochemistry. The expression of reactive oxygen species (ROS) and NOX activity were assessed by luciferase reporter gene assay, immunofluorescence analysis, and flow cytometry. Mutagenesis analysis, Alcian blue staining, reverse transcriptase polymerase chain reaction (RT-PCR), and western blot assay were used to determine whether p67phox was involved in suramin-enhanced chondrocyte phenotype maintenance.Aims
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Tendon is a bradytrophic and hypovascular tissue, hence, healing remains a major challenge. The molecular key events involved in successful repair have to be unravelled to develop novel strategies that reduce the risk of unfavourable outcomes such as non-healing, adhesion formation, and scarring. This review will consider the diverse pathophysiological features of tendon-derived cells that lead to failed healing, including misrouted differentiation (e.g. de- or transdifferentiation) and premature cell senescence, as well as the loss of functional progenitors. Many of these features can be attributed to disturbed cell-extracellular matrix (ECM) or unbalanced soluble mediators involving not only resident tendon cells, but also the cross-talk with immigrating immune cell populations. Unrestrained post-traumatic inflammation could hinder successful healing. Pro-angiogenic mediators trigger hypervascularization and lead to persistence of an immature repair tissue, which does not provide sufficient mechano-competence. Tendon repair tissue needs to achieve an ECM composition, structure, strength, and stiffness that resembles the undamaged highly hierarchically ordered tendon ECM. Adequate mechano-sensation and -transduction by tendon cells orchestrate ECM synthesis, stabilization by cross-linking, and remodelling as a prerequisite for the adaptation to the increased mechanical challenges during healing. Lastly, this review will discuss, from the cell biological point of view, possible optimization strategies for augmenting Achilles tendon (AT) healing outcomes, including adapted mechanostimulation and novel approaches by restraining neoangiogenesis, modifying stem cell niche parameters, tissue engineering, the modulation of the inflammatory cells, and the application of stimulatory factors. Cite this article:
