Objectives. This study aimed to explore the role of miR-320a in the pathogenesis of osteoarthritis (OA). Methods. Human cartilage cells (C28/I2) were transfected with miR-320a or antisense oligonucleotides (ASO)-miR-320a, and treated with
Objectives. The aim of this study was to investigate the role of miR-126 in the development of osteoarthritis, as well as the potential molecular mechanisms involved, in order to provide a theoretical basis for osteoarthritis treatment and a novel perspective for clinical therapy. Methods. Human chondrocyte cell line CHON-001 was administrated by different doses of interleukin (IL)-1β to simulate inflammation. Cell viability, migration, apoptosis, IL-6, IL-8, and tumour necrosis factor (TNF)-α expression, as well as expression of apoptosis-related factors, were measured to assess inflammation. miR-126 expression was measured by quantitative polymerase chain reaction (qPCR). Cells were then transfected with miR-126 inhibitor to assess the effect of miR-126 on IL-1β-injured CHON-001 cells. Expression of B-cell lymphoma 2 (Bcl-2) and the activity of mitogen-activated protein kinase (MAPK) / Jun N-terminal kinase (JNK) signaling pathway were measured by Western blot to explore the underlying mechanism through which miR-126 affects IL-1β-induced inflammation. Results. After
In order to clarify the role of cytokines in the remodelling of the grafted tendon for ligament reconstruction we compared the responses to interleukin (IL)-1β, platelet-derived growth factor (PDGF)-BB and transforming growth factor (TGF)-β1 of extrinsic fibroblasts infiltrating the frozen-thawed patellar tendon in rats with that of the normal tendon fibroblasts, in regard to the gene expression of matrix metalloproteinase (MMP)-13, using Northern blot analysis. We also examined, immunohistologically, the local expression of
Objective. Excessive mechanical stress on synovial joints causes osteoarthritis
(OA) and results in the production of prostaglandin E2 (PGE2), a
key molecule in arthritis, by synovial fibroblasts. However, the
relationship between arthritis-related molecules and mechanical
stress is still unclear. The purpose of this study was to examine
the synovial fibroblast response to cyclic mechanical stress using
an in vitro osteoarthritis model. Method. Human synovial fibroblasts were cultured on collagen scaffolds
to produce three-dimensional constructs. A cyclic compressive loading
of 40 kPa at 0.5 Hz was applied to the constructs, with or without
the administration of a cyclooxygenase-2 (COX-2) selective inhibitor
or dexamethasone, and then the concentrations of PGE2, interleukin-1β (IL-1β),
tumour necrosis factor-α (TNF-α), IL-6, IL-8 and COX-2 were measured. Results. The concentrations of PGE2, IL-6 and IL-8 in the loaded samples
were significantly higher than those of unloaded samples; however,
the concentrations of
Objectives. Osteoarthritis (OA) is characterised by articular cartilage degradation. MicroRNAs (miRNAs) have been identified in the development of OA. The purpose of our study was to explore the functional role and underlying mechanism of miR-138-5p in interleukin-1 beta (IL-1β)-induced extracellular matrix (ECM) degradation of OA cartilage. Materials and Methods. Human articular cartilage was obtained from patients with and without OA, and chondrocytes were isolated and stimulated by
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
Meniscus tears have been treated using partial meniscectomy to relieve pain in patients, although this leads to the onset of early osteoarthritis (OA). Cell-based therapies can help preserve the meniscus, although the presence of inflammatory cytokines compromises clinical outcomes. Anti-inflammatory drugs (e.g. celecoxib), can help to reduce pain in patients and in vitro studies suggest a beneficial effect on cytokine inhibited matrix content. Previously, we have demonstrated that the inhibitory effects of
While cell morphology has been recognized as a fundamental regulator of cell behavior, few studies have measured the complex cell morphological changes of chondrocytes using quantitative cell morphometry descriptors in relation to inflammation and phenotypic outcome. Acute vs. persistent exposure to
Previous research has shown catabolic cell signalling induced by TNF-α and
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 MMP-13 (p < 0.05) and IL1β (p < 0.001) was noticed. r-IR pretreatment of hNPCs cultured in pro-inflammatory conditions resulted in a rescue of metabolic activity (p < 0.001), as well as a decrease of
The purpose of this study was to evaluate the beneficial effects of r-Irisin (IR) on human primary tenocytes (hTCs) in vitro. Indeed, Irisin is secreted from muscles in response to exercise and mediates many beneficial effects on tissues and organs. Tissue samples (n=3) were analyzed by histology and immunohistochemistry for αVβ5 receptor. hTCs isolated, culture expanded were treated with: 1) RPMI medium as control; 2) IR at different concentrations; 3)
For patients who took joint replacement, one of the complications, aseptic joint loosening, could cause a high risk of revision surgery. Studies have shown that MSCs have the ability of homing and differentiating, and also have highly effective immune regulation and anti-inflammatory effects. However, few studies had focused on the stem cells in preventing the occurrence and development of aseptic loosening. In this research, we aimed to clarify whether human umbilical cord mesenchymal stem cells could inhibited the aseptic joint loosening caused by wear particles. A Cranial osteolysis mice model was established on mice to examine the effect of hUC-MSCs on the Titanium particles injection area through micro-CT. The amount of stem cells injected was 2 × 10 5 cells. One week later, the mouse Cranial were obtained for micro-CT scan, and then stained with HE analysis immunohistochemical analysis of TNF-α, CD68, CCL3 and
Introduction. Osteoarthritis (OA) is a prevalent joint disorder characterized by cartilage degeneration, inflammation, and pain. Current treatments provide only symptomatic relief, necessitating novel molecular targets. The caspase family, known for its roles in apoptosis and inflammation regulation, may additionally influence crucial processes for cartilage homeostasis such as differentiation and proliferation. However, the specific roles of individual caspases in OA pathogenesis remain unclear. This study aims to investigate the involvement of the caspase family in OA and as potential targets for therapy, with a focus on caspase-1 and -8. Method. Chondrocytes from both healthy and OA donors were cultured in 2D and 3D culture models and stimulated with TNF-α or
Intervertebral disc (IVD) degeneration is responsible for severe clinical symptoms including chronic back pain. Galectins are a family of carbohydrate-binding proteins, some of which can induce functional disease markers in IVD cells and other musculoskeletal diseases. Galectins −4 and −8 were shown to trigger disease-promoting activity in chondrocytes but their effects on IVD cells have not been investigated yet. This study elucidates the role of galectin-4 and −8 in IVD degeneration. Immunohistochemical evidence for the presence of galectin-4 and −8 in the IVD was comparatively provided in specimens of 36 patients with spondylochondrosis, spondylolisthesis, or spinal deformity. Confocal microscopy revealed co-localization of galectin-4 and −8 in chondrocyte clusters of degenerated cartilage. The immunohistochemical presence of galectin-4 correlated with histopathological and clinical degeneration scores of patients, whereas galectin-8 did not show significant correlations. The specimens were separated into annulus fibrosus (AF), nucleus pulposus (NP) and endplate, which was confirmed histologically. Separate cell cultures of AF and NP (n=20) were established and characterized using cell type-specific markers. Potential binding sites for galectins including sialylated N-glycans and LacdiNAc structures were determined in AF and NP cells using LC/ESI-MS-MS. To assess galectin functions, cell cultures were treated with recombinant galectin-4 or −8, in comparison to
Post-traumatic osteoarthritis (PTOA) is a subset of osteoarthritis, which occurs secondary to traumatic joint injury which is known to cause pathological changes to the osteochondral unit. Articular cartilage degradation is a primary hallmark of OA, and is normally associated with end-stage disease. However, subchondral bone marrow lesions are associated with joint injury, and may represent localized bone microdamage. Changes in the osteochondral unit have been traditionally studied using explant models, of which the femoral-head model is the most common. However, the bone damage caused during harvest can confound studies of microdamage. Thus, we used a novel patellar explant model to study osteochondral tissue dynamics and mechanistic changes in bone-cartilage crosstalk. Firstly, we characterized explants by comparing patella with femoral head models. Then, the patellar explants (n=269) were subjected to either mechanical or inflammatory stimulus. For mechanical stimulus 10% strain was applied at 0.5 and 1 Hz for 10 cycles. We also studied the responses of osteochondral tissues to 10ng/ml of TNF-α or
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
Aseptic loosening is a major cause of revision surgeries and occurs when osteolysis is stimulated around the implant by pro-inflammatory cytokines including
Osteoarthritis is a degenerative disease mainly caused by aging, although in younger patients (aged 25 – 50) it can be a consequence of sports-related injuries or trauma. This results in early osteoarthritis with subsequent changes in cartilage extracellular matrix. Cell-based tissue engineering approaches using mesenchymal stem cells (MSCs) are an ideal cell type for the treatment of early osteoarthritc defects. Our group has demonstrated in a clinical study, that interleukin-1β (IL-1β) was expressed in cartilage plugs from patients with early osteoarthritis. In vitro studies have shown that
Although 80% of fractures typically heal without any problems, there is a small proportion (<20%) that suffer complications such as delayed healing and potential progression to non-union. In patients with healing complications, the coordinated regulation between pro- and anti-inflammatory cytokines, such as interleukin-1β (IL-1β) and interleukin-1 receptor antagonist (IL-1Ra) respectively, is often dysregulated. The aim of this study is to develop a therapeutic strategy based on the local delivery of genes to reparative mesenchymal stromal cells (MSCs) migrating into the local fracture microenvironment, thereby promoting a more favourable healing environment to enhance fracture repair. Our approach involves the local delivery of nanoparticles complexing the non-viral vector polyethyleneimine (PEI) with therapeutic plasmid DNA (pDNA) encoding for IL-1Ra. pDNA encoding green fluorescent protein and Gaussia luciferase were used as reporter genes to determine the transfection efficiency of both rat and human MSCs using flow cytometry and to assess the transgene expression profile using a luciferase expression assay. The effect of transfection with PEI on the viability of MSCs was assessed using the metabolic assay Cell Titer Blue and dsDNA quantification. Levels of IL-1Ra produced by cells following transfection with nanoparticles encoding IL-1Ra was assessed using enzyme-linked immunosorbent assays (ELISA). HEK-Blue
Unresolved inflammatory processes in tendon healing have been related to the progression of tendinopathies. Thus, the management of tendon injuries may rely on cell-based strategies to identify and modulate tendon inflammatory cues. Pulsed electromagnetic field (PEMF) has been approved by FDA for orthopedics therapies and has been related to a reduction in pain and to improve healing. However, the influence of PEMF in tendon healing remains largely unknown. Human tendon resident cells (hTDCs) were cultured in an inflammatory environment induced by exogenous supplementation of
