Objectives. Studies which consider the molecular mechanisms of degeneration and regeneration of cartilaginous tissues are seriously hampered by problematic
Objectives. The aim of this study was to investigate the effect of hyperglycaemia on oxidative stress markers and inflammatory and matrix gene expression within tendons of normal and diabetic rats and to give insights into the processes involved in tendinopathy. Methods. Using tenocytes from normal Sprague-Dawley rats, cultured both in control and high glucose conditions, reactive oxygen species (ROS) production, cell proliferation, messenger
Objectives. Ligaments which heal spontaneously have a healing process that
is similar to skin wound healing. Menopause impairs skin wound healing
and may likewise impair ligament healing. Our purpose in this study
was to investigate the effect of surgical menopause on ligament
healing in a rabbit medial collateral ligament model. Methods. Surgical menopause was induced with ovariohysterectomy surgery
in adult female rabbits. Ligament injury was created by making a
surgical gap in the midsubstance of the medial collateral ligament.
Ligaments were allowed to heal for six or 14 weeks in the presence
or absence of oestrogen before being compared with uninjured ligaments. Molecular
assessment examined the messenger
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 IL-1β. The expression levels of miR-138-5p in cartilage and chondrocytes were both determined. After transfection with miR-138-5p mimics, allele-specific oligonucleotide (ASO)-miR-138-5p, or their negative controls, the messenger
Objectives. To investigate the appropriate dose and interval for the administration
of triamcinolone acetonide (TA) in treating tendinopathy to avoid
adverse effects such as tendon degeneration and rupture. Methods. Human rotator cuff-derived cells were cultured using three media:
regular medium (control), regular medium with 0.1 mg/mL of TA (low
TA group), and with 1.0 mg/mL of TA (high TA group). The cell morphology,
apoptosis, and viability were assessed at designated time points. Results. In the low TA group, the cells became flattened and polygonal
at seven days then returned to normal at 21 days. The cell apoptosis
ratio and messenger
We studied the presence of anabolic growth factors in human herniated intervertebral discs (IVD) using a reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry. Messenger
High-pressure lavage produces greater visible damage to bone at a macroscopic and microscopic level when compared with low-pressure lavage and can result in delay in the healing of fractures. Osteoblasts and adipocytes are derived from mesenchymal stem cells. Conditions which lead to bone loss often involve a switch from the osteoblast to adipocyte lineage. We have therefore examined the effect of high- and low-pressure irrigation on the differentiation of adipocytes. Calvaria-derived bone cells were exposed to either low-pressure or high-pressure irrigation with normal saline. After 14 days the cells were fixed and the osteoblasts and adipocytes quantified using Oil Red O to stain cytoplasmic lipid droplets (triglycerides) in the cells. Osteoblasts were quantified using a commercially available alkaline-phosphatase staining assay. A standard quantitative reverse transcription-polymerase chain reaction (RT-PCR) was performed. Messenger
We performed positron emission tomography (PET) with . 18. fluorine-fluoro-2-deoxy-D-glucose (FDG) on 55 patients with tumours involving the musculoskeletal system in order to evaluate its role in operative planning. The standardised uptake value (SUV) of FDG was calculated and, to distinguish malignancies from benign lesions, the cases were divided into high (≥ 1.9) and low (<
1.9) SUV groups. The sensitivity of PET for correctly diagnosing malignancy was 100% with a specificity of 76.9% and an overall accuracy of 83.0%. The mean SUV for metastatic lesions was twice that for primary sarcomas (p <
0.0015). Our results suggest that the SUV may be useful in differentiating malignant tumours from benign lesions. However, some of the latter, such as schwannomas, had high SUVs so that biopsy or wide resection was selected as the first operation. Thus, some other quantitative analysis may be required for preoperative planning in cases of high-SUV neurogenic benign tumours. The reverse transcription-polymerase chain reaction revealed that the
Adult mice lacking the transcription factor NFAT1 exhibit osteoarthritis (OA). The precise molecular mechanism for NFAT1 deficiency-induced osteoarthritic cartilage degradation remains to be clarified. This study aimed to investigate if NFAT1 protects articular cartilage (AC) against OA by directly regulating the transcription of specific catabolic and anabolic genes in articular chondrocytes. Through a combined approach of gene expression analysis and web-based searching of NFAT1 binding sequences, 25 candidate target genes that displayed aberrant expression in Objectives
Methods
Osteoporosis is a chronic disease. The aim of this study was to identify key genes in osteoporosis. Microarray data sets GSE56815 and GSE56814, comprising 67 osteoporosis blood samples and 62 control blood samples, were obtained from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) were identified in osteoporosis using Limma package (3.2.1) and Meta-MA packages. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses were performed to identify biological functions. Furthermore, the transcriptional regulatory network was established between the top 20 DEGs and transcriptional factors using the UCSC ENCODE Genome Browser. Receiver operating characteristic (ROC) analysis was applied to investigate the diagnostic value of several DEGs.Objectives
Methods
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. 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.Objectives
Methods
The role of mechanical stress and transforming growth factor beta 1 (TGF-β1) is important in the initiation and progression of osteoarthritis (OA). However, the underlying molecular mechanisms are not clearly known. In this study, TGF-β1 from osteoclasts and knee joints were analyzed using a co-cultured cell model and an OA rat model, respectively. Five patients with a femoral neck fracture (four female and one male, mean 73.4 years (68 to 79)) were recruited between January 2015 and December 2015. Results showed that TGF-β1 was significantly upregulated in osteoclasts by cyclic loading in a time- and dose-dependent mode. The osteoclasts were subjected to cyclic loading before being co-cultured with chondrocytes for 24 hours.Objectives
Methods
Platelet-rich fibrin matrix (PRFM) has been proved to enhance tenocyte proliferation but has mixed results when used during rotator cuff repair. The optimal PRFM preparation protocol should be determined before clinical application. To screen the best PRFM to each individual’s tenocytes effectively, small-diameter culture wells should be used to increase variables. The gelling effect of PRFM will occur when small-diameter culture wells are used. A co-culture device should be designed to avoid this effect. Tenocytes harvested during rotator cuff repair and blood from a healthy volunteer were used. Tenocytes were seeded in 96-, 24-, 12-, and six-well plates and co-culture devices. Appropriate volumes of PRFM, according to the surface area of each culture well, were treated with tenocytes for seven days. The co-culture device was designed to avoid the gelling effect that occurred in the small-diameter culture well. Cell proliferation was analyzed by water soluble tetrazolium-1 (WST-1) bioassay.Objectives
Methods
This study looked to analyse the expression levels of microRNA-140-3p and microRNA-140-5p in synovial fluid, and their correlations to the severity of disease regarding knee osteoarthritis (OA). Knee joint synovial fluid samples were collected from 45 patients with OA of the knee (15 mild, 15 moderate and 15 severe), ten healthy volunteers, ten patients with gouty arthritis, and ten with rheumatoid arthritis. The Kellgren–Lawrence grading (KLG) was used to assess the radiological severity of knee OA, and the patients were stratified into mild (KLG < 2), moderate (KLG = 2), and severe (KLG > 2). The expression of miR-140-3p and miR-140-5p of individual samples was measured by SYBR Green quantitative polymerase chain reaction (PCR) analysis. The expression of miR-140-3p and miR-140-5p was normalised to U6 internal control using the 2-△△CT method. All data were processed using SPSS software.Objectives
Methods
The objective of this study was to investigate the therapeutic effect of peripheral blood mononuclear cells (PBMNCs) treated with quality and quantity control culture (QQ-culture) to expand and fortify angiogenic cells on the acceleration of fracture healing. Human PBMNCs were cultured for seven days with the QQ-culture method using a serum-free medium containing five specific cytokines and growth factors. The QQ-cultured PBMNCs (QQMNCs) obtained were counted and characterised by flow cytometry and real-time polymerase chain reaction (RT-PCR). Angiogenic and osteo-inductive potentials were evaluated using tube formation assays and co-culture with mesenchymal stem cells with osteo-inductive medium Objectives
Methods
Regenerative medicine is an emerging field aimed at the repair and regeneration of various tissues. To this end, cytokines (CKs), growth factors (GFs), and stem/progenitor cells have been applied in this field. However, obtaining and preparing these candidates requires invasive, costly, and time-consuming procedures. We hypothesised that skeletal muscle could be a favorable candidate tissue for the concept of a point-of-care approach. The purpose of this study was to characterize and confirm the biological potential of skeletal muscle supernatant for use in regenerative medicine. Semitendinosus muscle was used after harvesting tendon from patients who underwent anterior cruciate ligament reconstructions. A total of 500 milligrams of stripped muscle was minced and mixed with 1 mL of saline. The collected supernatant was analysed by enzyme-linked immunosorbent assay (ELISA) and flow cytometry. The biological effects of the supernatant on cell proliferation, osteogenesis, and angiogenesis in vitro were evaluated using human mesenchymal stem cells (hMSCs) and human umbilical cord vein endothelial cells (HUVECs).Objectives
Methods
This study aimed to explore the role of miR-320a in the pathogenesis of osteoarthritis (OA). Human cartilage cells (C28/I2) were transfected with miR-320a or antisense oligonucleotides (ASO)-miR-320a, and treated with IL-1β. Subsequently the expression of collagen type II alpha 1 (Col2α1) and aggrecan (ACAN), and the concentrations of sulfated glycosaminoglycans (sGAG) and matrix metallopeptidase 13 (MMP-13), were assessed. Luciferase reporter assay, qRT-PCR, and Western blot were performed to explore whether pre-B-cell leukemia Homeobox 3 (PBX3) was a target of miR-320a. Furthermore, cells were co-transfected with miR-320a and PBX3 expressing vector, or cells were transfected with miR-320a and treated with a nuclear factor kappa B (NF-κB) antagonist MG132. The changes in Col2α1 and ACAN expression, and in sGAG and MMP-13 concentrations, were measured again. Statistical comparisons were made between two groups by using the two-tailed paired Objectives
Methods
The molecular mechanism of rheumatoid arthritis (RA) remains elusive. We conducted a protein-protein interaction network-based integrative analysis of genome-wide association studies (GWAS) and gene expression profiles of RA. We first performed a dense search of RA-associated gene modules by integrating a large GWAS meta-analysis dataset (containing 5539 RA patients and 20 169 healthy controls), protein interaction network and gene expression profiles of RA synovium and peripheral blood mononuclear cells (PBMCs). Gene ontology (GO) enrichment analysis was conducted by DAVID. The protein association networks of gene modules were generated by STRING.Objectives
Methods
Triamcinolone acetonide (TA) is widely used for the treatment of rotator cuff injury because of its anti-inflammatory properties. However, TA can also produce deleterious effects such as tendon degeneration or rupture. These harmful effects could be prevented by the addition of platelet-rich plasma (PRP), however, the anti-inflammatory and anti-degenerative effects of the combined use of TA and PRP have not yet been made clear. The objective of this study was to determine how the combination of TA and PRP might influence the inflammation and degeneration of the rotator cuff by examining rotator cuff-derived cells induced by interleukin (IL)-1ß. Rotator cuff-derived cells were seeded under inflammatory stimulation conditions (with serum-free medium with 1 ng/ml IL-1ß for three hours), and then cultured in different media: serum-free (control group), serum-free + TA (0.1mg/ml) (TA group), serum-free + 10% PRP (PRP group), and serum-free + TA (0.1mg/ml) + 10% PRP (TA+PRP group). Cell morphology, cell viability, and expression of inflammatory and degenerative mediators were assessed.Objectives
Methods
To assess the structure and extracellular matrix molecule expression of osteogenic cell sheets created via culture in medium with both dexamethasone (Dex) and ascorbic acid phosphate (AscP) compared either Dex or AscP alone. Osteogenic cell sheets were prepared by culturing rat bone marrow stromal cells in a minimal essential medium (MEM), MEM with AscP, MEM with Dex, and MEM with Dex and AscP (Dex/AscP). The cell number and messenger (m)RNA expression were assessed Objectives
Methods
We have observed clinical cases where bone is formed in the overlaying muscle covering surgically created bone defects treated with a hydroxyapatite/calcium sulphate biomaterial. Our objective was to investigate the osteoinductive potential of the biomaterial and to determine if growth factors secreted from local bone cells induce osteoblastic differentiation of muscle cells. We seeded mouse skeletal muscle cells C2C12 on the hydroxyapatite/calcium sulphate biomaterial and the phenotype of the cells was analysed. To mimic surgical conditions with leakage of extra cellular matrix (ECM) proteins and growth factors, we cultured rat bone cells ROS 17/2.8 in a bioreactor and harvested the secreted proteins. The secretome was added to rat muscle cells L6. The phenotype of the muscle cells after treatment with the media was assessed using immunostaining and light microscopy.Objectives
Materials and Methods
Osteoporosis is a systemic skeletal disorder characterized by reduced bone mass and deterioration of bone microarchitecture, which results in increased bone fragility and fracture risk. Casein kinase 2-interacting protein-1 (CKIP-1) is a protein that plays an important role in regulation of bone formation. The effect of CKIP-1 on bone formation is mainly mediated through negative regulation of the bone morphogenetic protein pathway. In addition, CKIP-1 has an important role in the progression of osteoporosis. This review provides a summary of the recent studies on the role of CKIP-1 in osteoporosis development and treatment.
This study aimed to examine the effects of SRT1720, a potent SIRT1 activator, on osteoarthritis (OA) progression using an experimental OA model. Osteoarthritis was surgically induced by destabilization of the medial meniscus in eight-week-old C57BL/6 male mice. SRT1720 was administered intraperitoneally twice a week after surgery. Osteoarthritis progression was evaluated histologically using the Osteoarthritis Research Society International (OARSI) score at four, eight, 12 and 16 weeks. The expression of SIRT1, matrix metalloproteinase 13 (MMP-13), a disintegrin and metalloproteinase with thrombospondin motifs-5 (ADAMTS-5), cleaved caspase-3, PARP p85, and acetylated nuclear factor (NF)-κB p65 in cartilage was examined by immunohistochemistry. Synovitis was also evaluated histologically. Primary mouse epiphyseal chondrocytes were treated with SRT1720 in the presence or absence of interleukin 1 beta (IL-1β), and gene expression changes were examined by real-time polymerase chain reaction (PCR).Objectives
Methods
Interleukin 18 (IL-18) is a regulatory cytokine that degrades the disc matrix. Bone morphogenetic protein-2 (BMP-2) stimulates synthesis of the disc extracellular matrix. However, the combined effects of BMP-2 and IL-18 on human intervertebral disc degeneration have not previously been reported. The aim of this study was to investigate the effects of the anabolic cytokine BMP-2 and the catabolic cytokine IL-18 on human nucleus pulposus (NP) and annulus fibrosus (AF) cells and, therefore, to identify potential therapeutic and clinical benefits of recombinant human (rh)BMP-2 in intervertebral disc degeneration. Levels of IL-18 were measured in the blood of patients with intervertebral disc degenerative disease and in control patients. Human NP and AF cells were cultured in a NP cell medium and treated with IL-18 or IL-18 plus BMP-2. mRNA levels of target genes were measured by real-time polymerase chain reaction, and protein levels of aggrecan, type II collagen, SOX6, and matrix metalloproteinase 13 (MMP13) were assessed by western blot analysis.Objectives
Methods
The cytotoxicity induced by cobalt ions (Co2+) and cobalt nanoparticles (Co-NPs) which released following the insertion of a total hip prosthesis, has been reported. However, little is known about the underlying mechanisms. In this study, we investigate the toxic effect of Co2+ and Co-NPs on liver cells, and explain further the potential mechanisms. Co-NPs were characterised for size, shape, elemental analysis, and hydrodynamic diameter, and were assessed by Transmission Electron Microscope, Scanning Electron Microscope, Energy Dispersive X-ray Spectroscopy and Dynamic Light Scattering. BRL-3A cells were used in this study. Cytotoxicity was evaluated by MTT and lactate dehydrogenase release assay. In order to clarify the potential mechanisms, reactive oxygen species, Bax/Bcl-2 mRNA expression, IL-8 mRNA expression and DNA damage were assessed on BRL-3A cells after Co2+ or Co-NPs treatment.Objectives
Methods
Ubiquitin E3 ligase-mediated protein degradation regulates osteoblast function. Itch, an E3 ligase, affects numerous cell functions by regulating ubiquitination and proteasomal degradation of related proteins. However, the Itch-related cellular and molecular mechanisms by which osteoblast differentiation and function are elevated during bone fracture repair are as yet unknown. We examined the expression levels of E3 ligases and NF-κB members in callus samples during bone fracture repair by quantitative polymerase chain reaction (qPCR) and the total amount of ubiquitinated proteins by Western blot analysis in wild-type (WT) mice. The expression levels of osteoblast-associated genes in fracture callus from Itch knockout (KO) mice and their WT littermates were examined by qPCR. The effect of NF-κB on Itch expression in C2C12 osteoblast cells was determined by a chromatin immunoprecipitation (ChIP) assay.Objectives
Methods
The present study describes a novel technique for revitalising allogenic intrasynovial tendons by combining cell-based therapy and mechanical stimulation in an Specifically, canine flexor digitorum profundus tendons were used for this study and were divided into the following groups: (1) untreated, unprocessed normal tendon; (2) decellularised tendon; (3) bone marrow stromal cell (BMSC)-seeded tendon; and (4) BMSC-seeded and cyclically stretched tendon. Lateral slits were introduced on the tendon to facilitate cell seeding. Tendons from all four study groups were distracted by a servohydraulic testing machine. Tensile force and displacement data were continuously recorded at a sample rate of 20 Hz until 200 Newton of force was reached. Before testing, the cross-sectional dimensions of each tendon were measured with a digital caliper. Young’s modulus was calculated from the slope of the linear region of the stress-strain curve. The BMSCs were labeled for histological and cell viability evaluation on the decellularized tendon scaffold under a confocal microscope. Gene expression levels of selected extracellular matrix tendon growth factor genes were measured. Results were reported as mean ± SD and data was analyzed with one-way ANOVAs followed by Tukey’s post hoc multiple-comparison test.Objectives
Methods
Osteophytes are products of active endochondral and intramembranous ossification, and therefore could theoretically provide significant efficacy as bone grafts. In this study, we compared the bone mineralisation effectiveness of osteophytes and cancellous bone, including their effects on secretion of growth factors and anabolic effects on osteoblasts. Osteophytes and cancellous bone obtained from human patients were transplanted onto the calvaria of severe combined immunodeficient mice, with Calcein administered intra-peritoneally for fluorescent labelling of bone mineralisation. Conditioned media were prepared using osteophytes and cancellous bone, and growth factor concentration and effects of each graft on proliferation, differentiation and migration of osteoblastic cells were assessed using enzyme-linked immunosorbent assays, MTS ((3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium)) assays, quantitative real-time polymerase chain reaction, and migration assays.Objectives
Methods
After an injury, the biological reattachment of tendon to bone is a challenge because healing takes place between a soft (tendon) and a hard (bone) tissue. Even after healing, the transition zone in the enthesis is not completely regenerated, making it susceptible to re-injury. In this study, we aimed to regenerate Achilles tendon entheses (ATEs) in wounded rats using a combination of kartogenin (KGN) and platelet-rich plasma (PRP). Wounds created in rat ATEs were given three different treatments: kartogenin platelet-rich plasma (KGN-PRP); PRP; or saline (control), followed by histological and immunochemical analyses, and mechanical testing of the rat ATEs after three months of healing.Objectives
Methods
Animal models have been developed that allow simulation of post-traumatic joint contracture. One such model involves contracture-forming surgery followed by surgical capsular release. This model allows testing of antifibrotic agents, such as rosiglitazone. A total of 20 rabbits underwent contracture-forming surgery. Eight weeks later, the animals underwent a surgical capsular release. Ten animals received rosiglitazone (intramuscular initially, then orally). The animals were sacrificed following 16 weeks of free cage mobilisation. The joints were tested biomechanically, and the posterior capsule was assessed histologically and via genetic microarray analysis.Aims
Methods
This study aimed to investigate the functional effects of microRNA (miR)-214-5p on osteoblastic cells, which might provide a potential role of miR-214-5p in bone fracture healing. Blood samples were obtained from patients with hand fracture or intra-articular calcaneal fracture and from healthy controls (HCs). Expression of miR-214-5p was monitored by qRT-PCR at day 7, 14 and 21 post-surgery. Mouse osteoblastic MC3T3-E1 cells were transfected with antisense oligonucleotides (ASO)-miR-214-5p, collagen type IV alpha 1 (COL4A1) vector or their controls; thereafter, cell viability, apoptotic rate, and the expression of collagen type I alpha 1 (COL1A1), type II collagen (COL-II), and type X collagen (COL-X) were determined. Luciferase reporter assay, qRT-PCR, and Western blot were performed to ascertain whether COL4A1 was a target of miR-214-5p.Objectives
Methods
Recent studies have shown that systemic injection of rapamycin can prevent the development of osteoarthritis (OA)-like changes in human chondrocytes and reduce the severity of experimental OA. However, the systemic injection of rapamycin leads to many side effects. The purpose of this study was to determine the effects of intra-articular injection of Torin 1, which as a specific inhibitor of mTOR which can cause induction of autophagy, is similar to rapamycin, on articular cartilage degeneration in a rabbit osteoarthritis model and to investigate the mechanism of Torin 1’s effects on experimental OA. Collagenase (type II) was injected twice into both knees of three-month-old rabbits to induce OA, combined with two intra–articular injections of Torin 1 (400 nM). Degeneration of articular cartilage was evaluated by histology using the Mankin scoring system at eight weeks after injection. Chondrocyte degeneration and autophagosomes were observed by transmission electron microscopy. Matrix metallopeptidase-13 (MMP-13) and vascular endothelial growth factor (VEGF) expression were analysed by quantitative RT-PCR (qPCR).Beclin-1 and light chain 3 (LC3) expression were examined by Western blotting.Objectives
Methods
Sustained intra-articular delivery of pharmacological agents is an attractive modality but requires use of a safe carrier that would not induce cartilage damage or fibrosis. Collagen scaffolds are widely available and could be used intra-articularly, but no investigation has looked at the safety of collagen scaffolds within synovial joints. The aim of this study was to determine the safety of collagen scaffold implantation in a validated A total of 96 rabbits were randomly and equally assigned to four different groups: arthrotomy alone; arthrotomy and collagen scaffold placement; contracture surgery; and contracture surgery and collagen scaffold placement. Animals were killed in equal numbers at 72 hours, two weeks, eight weeks, and 24 weeks. Joint contracture was measured, and cartilage and synovial samples underwent histological analysis.Objectives
Materials and Methods
Intermittently administered parathyroid hormone (PTH 1-34) has been shown to promote bone formation in both human and animal studies. The hormone and its analogues stimulate both bone formation and resorption, and as such at low doses are now in clinical use for the treatment of severe osteoporosis. By varying the duration of exposure, parathyroid hormone can modulate genes leading to increased bone formation within a so-called ‘anabolic window’. The osteogenic mechanisms involved are multiple, affecting the stimulation of osteoprogenitor cells, osteoblasts, osteocytes and the stem cell niche, and ultimately leading to increased osteoblast activation, reduced osteoblast apoptosis, upregulation of Wnt/β-catenin signalling, increased stem cell mobilisation, and mediation of the RANKL/OPG pathway. Ongoing investigation into their effect on bone formation through ‘coupled’ and ‘uncoupled’ mechanisms further underlines the impact of intermittent PTH on both cortical and cancellous bone. Given the principally catabolic actions of continuous PTH, this article reviews the skeletal actions of intermittent PTH 1-34 and the mechanisms underlying its effect.
In order to screen the altered gene expression profile in peripheral blood mononuclear cells of patients with osteoporosis, we performed an integrated analysis of the online microarray studies of osteoporosis. We searched the Gene Expression Omnibus (GEO) database for microarray studies of peripheral blood mononuclear cells in patients with osteoporosis. Subsequently, we integrated gene expression data sets from multiple microarray studies to obtain differentially expressed genes (DEGs) between patients with osteoporosis and normal controls. Gene function analysis was performed to uncover the functions of identified DEGs.Objectives
Methods
Rotator cuff tears are among the most frequent upper extremity injuries. Current treatment strategies do not address the poor quality of the muscle and tendon following chronic rotator cuff tears. Hypoxia-inducible factor-1 alpha (HIF-1α) is a transcription factor that activates many genes that are important in skeletal muscle regeneration. HIF-1α is inhibited under normal physiological conditions by the HIF prolyl 4-hydroxylases (PHDs). In this study, we used a pharmacological PHD inhibitor, GSK1120360A, to enhance the activity of HIF-1α following the repair of a chronic cuff tear, and measured muscle fibre contractility, fibrosis, gene expression, and enthesis mechanics. Chronic supraspinatus tears were induced in adult rats, and repaired 28 days later. Rats received 0 mg/kg, 3 mg/kg, or 10 mg/kg GSK1120360A daily. Collagen content, contractility, fibre type distribution and size, the expression of genes involved in fibrosis, lipid accumulation, atrophy and inflammation, and the mechanical properties of the enthesis were then assessed two weeks following surgical repair.Objectives
Methods
Rotator cuff tears are among the most common and debilitating
upper extremity injuries. Chronic cuff tears result in atrophy and
an infiltration of fat into the muscle, a condition commonly referred
to as ‘fatty degeneration’. While stem cell therapies hold promise
for the treatment of cuff tears, a suitable immunodeficient animal
model that could be used to study human or other xenograft-based
therapies for the treatment of rotator cuff injuries had not previously
been identified. A full-thickness, massive supraspinatus and infraspinatus tear
was induced in adult T-cell deficient rats. We hypothesised that,
compared with controls, 28 days after inducing a tear we would observe
a decrease in muscle force production, an accumulation of type IIB
fibres, and an upregulation in the expression of genes involved
with muscle atrophy, fibrosis and inflammation.Objectives
Methods
Venous thromboembolism (VTE) is a major potential complication following orthopaedic surgery. Subcutaneously administered enoxaparin has been used as the benchmark to reduce the incidence of VTE. However, concerns have been raised regarding the long-term administration of enoxaparin and its possible negative effects on bone healing and bone density with an increase of the risk of osteoporotic fractures. New oral anticoagulants such as rivaroxaban have recently been introduced, however, there is a lack of information regarding how these drugs affect bone metabolism and post-operative bone healing. We measured the migration and proliferation capacity of mesenchymal stem cells (MSCs) under enoxaparin or rivaroxaban treatment for three consecutive weeks, and evaluated effects on MSC mRNA expression of markers for stress and osteogenic differentiation.Objectives
Methods
Resveratrol is a polyphenolic compound commonly found in the
skins of red grapes. Sirtuin 1 (SIRT1) is a human gene that is activated
by resveratrol and has been shown to promote longevity and boost
mitochondrial metabolism. We examined the effect of resveratrol
on normal and osteoarthritic (OA) human chondrocytes. Normal and OA chondrocytes were incubated with various concentrations
of resveratrol (1 µM, 10 µM, 25 µM and 50 µM) and cultured for 24,
48 or 72 hours or for six weeks. Cell proliferation, gene expression,
and senescence were evaluated.Background
Methods
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 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.Objective
Method
The role of inflammatory cells and their products in tendinopathy is not completely understood. Pro-inflammatory cytokines are upregulated after oxidative and other forms of stress. Based on observations that increased cytokine expression has been demonstrated in cyclically-loaded tendon cells we hypothesised that because of their role in oxidative stress and apoptosis, pro-inflammatory cytokines may be present in rodent and human models of tendinopathy. A rat supraspinatus tendinopathy model produced by running overuse was investigated at the genetic level by custom micro-arrays. Additionally, samples of torn supraspinatus tendon and matched intact subscapularis tendon were collected from patients undergoing arthroscopic shoulder surgery for rotator-cuff tears and control samples of subscapularis tendon from ten patients with normal rotator cuffs undergoing arthroscopic stabilisation of the shoulder were also obtained. These were all evaluated using semiquantitative reverse transcription polymerase chain-reaction and immunohistochemistry. We identified significant upregulation of pro-inflammatory cytokines and apoptotic genes in the rodent model (p = 0.005). We further confirmed significantly increased levels of cytokine and apoptotic genes in human supraspinatus and subscapularis tendon harvested from patients with rotator cuff tears (p = 0.0008). These findings suggest that pro-inflammatory cytokines may play a role in tendinopathy and may provide a target for preventing tendinopathies.
The patellofemoral joint is an important source of symptoms in osteoarthritis of the knee. We have used a newly designed surgical model of patellar strengthening to induce osteoarthritis in BALB/c mice and to establish markers by investigating the relationship between osteoarthritis and synovial levels of matrix metalloproteinases (MMPs). Osteoarthritis was induced by using this microsurgical technique under direct vision without involving the cavity of the knee. Degeneration of cartilage was assessed by the Mankin score and synovial tissue was used to determine the mRNA expression levels of MMPs. Irrigation fluid from the knee was used to measure the concentrations of MMP-3 and MMP-9. Analysis of cartilage degeneration was correlated with the levels of expression of MMP. After operation the patellofemoral joint showed evidence of mild osteoarthritis at eight weeks and further degenerative changes by 12 weeks. The level of synovial MMP-9 mRNA correlated with the Mankin score at eight weeks, but not at 12 weeks. The levels of MMP-2, MMP-3 and MMP-14 mRNA correlated with the Mankin score at 12 weeks. An increase in MMP-3 was observed from four weeks up to 16 weeks. MMP-9 was notably increased at eight weeks, but the concentration at 16 weeks had decreased to the level observed at four weeks. Our observations suggest that MMP-2, MMP-3 and MMP-14 could be used as markers of the progression of osteoarthritic change.
The aim of this study was to evaluate the cultivation potential of cartilage taken from the debrided edge of a chronic lesion of the articular surface. A total of 14 patients underwent arthroscopy of the knee for a chronic lesion on the femoral condyles or trochlea. In addition to the routine cartilage biopsy, a second biopsy of cartilage was taken from the edge of the lesion. The cells isolated from both sources underwent parallel cultivation as monolayer and three-dimensional (3D) alginate culture. The cell yield, viability, capacity for proliferation, morphology and the expressions of typical cartilage genes (collagen I, COL1; collagen II, COL2; aggrecan, AGR; and versican, VER) were assessed. The cartilage differentiation indices (COL2/COL1, AGR/VER) were calculated. The control biopsies revealed a higher mean cell yield (1346 cells/mg Our results suggest that the cultivation of chondrocytes solely from the edges of the lesion cannot be recommended for use in autologous chondrocyte implantation.
The haematoma occurring at the site of a fracture is known to play an important role in bone healing. We have recently shown the presence of progenitor cells in human fracture haematoma and demonstrated that they have the capacity for multilineage mesenchymal differentiation. There have been many studies which have shown that low-intensity pulsed ultrasound (LIPUS) stimulates the differentiation of a variety of cells, but none has investigated the effects of LIPUS on cells derived from human fracture tissue including human fracture haematoma-derived progenitor cells (HCs). In this
We isolated multilineage mesenchymal progenitor cells from haematomas collected from fracture sites. After the haematoma was manually removed from the fracture site it was cut into strips and cultured. Homogenous fibroblastic adherent cells were obtained. Flow cytometry revealed that the adherent cells were consistently positive for mesenchymal stem-cell-related markers CD29, CD44, CD105 and CD166, and were negative for the haemopoietic markers CD14, CD34, CD45 and CD133 similar to bone-marrow-derived mesenchymal stem cells. In the presence of lineage-specific induction factors the adherent cells could differentiate Our results indicate that haematomas found at a fracture site contain multilineage mesenchymal progenitor cells and play an important role in bone healing. Our findings imply that to enhance healing the haematoma should not be removed from the fracture site during osteosynthesis.
Gene therapy with insulin-like growth factor-1 (IGF-1) increases matrix production and enhances chondrocyte proliferation and survival A total of 16 horses underwent arthroscopic repair of a single 15 mm cartilage defect in each femoropatellar joint. One joint received 2 × 107 AdIGF-1 modified chondrocytes and the contralateral joint received 2 × 107 naive (unmodified) chondrocytes. Repairs were analysed at four weeks, nine weeks and eight months after surgery. Morphological and histological appearance, IGF-1 and collagen type II gene expression (polymerase chain reaction, Genetic modification of chondrocytes significantly increased IGF-1 mRNA and ligand production in repair tissue for up to nine weeks following transplantation. The gross and histological appearance of IGF-1 modified repair tissue was improved over control defects. Gross filling of defects was significantly improved at four weeks, and a more hyaline-like tissue covered the lesions at eight months. Histological outcome at four and nine weeks post-transplantation revealed greater tissue filling of defects transplanted with genetically modified chondrocytes, whereas repair tissue in control defects was thin and irregular and more fibrous. Collagen type II expression in IGF-1 gene-transduced defects was increased 100-fold at four weeks and correlated with increased collagen type II immunoreaction up to eight months. Genetic modification of chondrocytes with AdIGF-1 prior to transplantation improved early (four to nine weeks), and to a lesser degree long-term, cartilage healing in the equine model. The equine model of cartilage healing closely resembles human clinical cartilage repair. The results of this study suggest that cartilage healing can be enhanced through genetic modification of chondrocytes prior to transplantation.
The gelatin-based haemostyptic compound Spongostan was tested as a three-dimensional (3D) chondrocyte matrix in an in vitro model for autologous chondrocyte transplantation using cells harvested from bovine knees. In a control experiment of monolayer cultures, the proliferation or de-differentiation of bovine chondrocytes was either not or only marginally influenced by the presence of Spongostan (0.3 mg/ml). In monolayers and 3-D Minusheet culture chambers, the cartilage-specific differentiation markers aggrecan and type-II collagen were ubiquitously present in a cell-associated fashion and in the pericellular matrix. The Minusheet cultures usually showed a markedly higher mRNA expression than monolayer cultures irrespective of whether Spongostan had been present or not during culture. Although the de-differentiation marker type-I collagen was also present, the ratio of type-I to type-II collagen or aggrecan to type-I collagen remained higher in Minusheet 3-D cultures than in monolayer cultures irrespective of whether Spongostan had been included in or excluded from the monolayer cultures. The concentration of GAG in Minusheet cultures reached its maximum after 14 days with a mean of 0.83 ± 0.8 μg/106 cells; mean ±, Our results suggest that Spongostan is in principle suitable as a 3-D chondrocyte matrix, as demonstrated in Minusheet chambers, in particular for a culture period of 14 days. Clinically, differentiating effects on chondrocytes, simple handling and optimal formability may render Spongostan an attractive 3-D scaffold for autologous chondrocyte transplantation.
We have investigated whether cells derived from haemarthrosis caused by injury to the anterior cruciate ligament could differentiate into the osteoblast lineage Our results suggest that the haemarthrosis induced by injury to the anterior cruciate ligament contains osteoprogenitor cells and is a potential alternative source for cell-based treatment in such injury.
The aim of our study was to investigate the effect of platelet-rich plasma on the proliferation and differentiation of rat bone-marrow cells and to determine an optimal platelet concentration in plasma for osseous tissue engineering. Rat bone-marrow cells embedded in different concentrations of platelet-rich plasma gel were cultured for six days. Their potential for proliferation and osteogenic differentiation was analysed. Using a rat limb-lengthening model, the cultured rat bone-marrow cells with platelet-rich plasma of variable concentrations were transplanted into the distraction gap and the quality of the regenerate bone was evaluated radiologically. Cellular proliferation was enhanced in all the platelet-rich plasma groups in a dose-dependent manner. Although no significant differences in the production and mRNA expression of alkaline phosphatase were detected among these groups, mature bone regenerates were more prevalent in the group with the highest concentration of platelets. Our results indicate that a high platelet concentration in the platelet-rich plasma in combination with osteoblastic cells could accelerate the formation of new bone during limb-lengthening procedures.
We investigated the effect of progesterone on the nerve during lengthening of the limb in rats. The sciatic nerves of rats were elongated by leg lengthening for ten days at 3 mm per day. On alternate days between the day after the operation and nerve dissection, the progesterone-treated group received subcutaneous injections of 1 mg progesterone in sesame oil and the control group received oil only. On the fifth, tenth and 17th day, the sciatic nerves were excised at the midpoint of the femur and the mRNA expression level of myelin protein P0 was analysed by quantitative real time polymerase chain reaction. On day 52 nodal length was examined by electron microscopy, followed by an examination of the compound muscle action potential (C-MAP) amplitude and the motor conduction velocity (MCV) of the tibial nerve on days 17 and 52. The P0 (a major myelin glycoprotein) mRNA expression level in the progesterone-treated group increased by 46.6% and 38.7% on days five and ten, respectively. On day 52, the nodal length in the progesterone-treated group was smaller than that in the control group, and the MCV of the progesterone-treated group had been restored to normal. Progesterone might accelerate the restoration of demyelination caused by nerve elongation by activating myelin synthesis.