Multiple biochemical biomarkers have been previously investigated for the diagnosis, prognosis and response to treatment of articular cartilage damage, including osteoarthritis (OA). Synovial fluid (SF) biomarker measurement is a potential method to predict treatment response and effectiveness. However, the significance of different biomarkers and their correlation to clinical outcomes remains unclear. This systematic review evaluated current SF biomarkers used in investigation of cartilage degeneration or regeneration in the knee joint and correlated these biomarkers with clinical outcomes following cartilage repair or regeneration interventions. PubMed, Institute of Science Index, Scopus, Cochrane Central Register of Controlled Trials, and Embase databases were searched. Studies evaluating SF biomarkers and clinical outcomes following cartilage repair intervention were included. Two researchers independently performed data extraction and QUADAS-2 analysis. Biomarker inclusion, change following intervention and correlation with clinical outcome was compared. 9 studies were included. Study heterogeneity precluded meta-analysis. There was significant variation in sampling and analysis. 33 biomarkers were evaluated in addition to microRNA and catabolic/anabolic ratios. Five studies reported on correlation of biomarkers with six biomarkers significantly correlated with clinical outcomes following intervention. However, correlation was only demonstrated in isolated studies. This review demonstrates significant difficulties in drawing conclusions regarding the importance of SF biomarkers based on the available literature. Improved standardisation for collection and analysis of SF samples is required. Future publications should also focus on clinical outcome scores and seek to correlate biomarkers with progression to further understand the significance of identified markers in a clinical context.
Dislocation is one of the most common complications in total hip arthroplasty (THA) and is primarily driven by bony or prosthetic impingement. The aim of this study was two-fold. First, to develop a simulation that incorporates the functional position of the femur and pelvis and instantaneously determines range of motion (ROM) limits. Second, to assess the number of patients for whom their functional bony alignment escalates impingement risk. 468 patients underwent a preoperative THA planning protocol that included functional x-rays and a lower limb CT scan. The CT scan was segmented and landmarked, and the x-rays were measured for pelvic tilt, femoral rotation, and preoperative leg length discrepancy (LLD). All patients received 3D templating with the same implant combination (Depuy; Corail/Pinnacle). Implants were positioned according to standardised criteria. Each patient was simulated in a novel ROM simulation that instantaneously calculates bony and prosthetic impingement limits in functional movements. Simulated motions included flexion and standing-external rotation (ER). Each patient's ROM was simulated with their bones oriented in both functional and neutral positions. 13% patients suffered a ROM impingement for functional but not neutral extension-ER. As a result, 48% patients who failed the functional-ER simulation would not be detected without consideration of the functional bony alignment. 16% patients suffered a ROM impingement for functional but not neutral flexion. As a result, 65% patients who failed the flexion simulation would not be detected without consideration of the functional bony alignment. We have developed a ROM simulation for use with preoperative planning for THA surgery that can solve bony and prosthetic impingement limits instantaneously. The advantage of our ROM simulation over previous simulations is instantaneous impingement detection, not requiring implant geometries to be analysed prior to use, and addressing the functional position of both the femur and pelvis.
In 2021, Vigdorchik et al. published a large multicentre study validating their simple Hip-Spine Classification for determining patient-specific acetabular component positioning in total hip arthroplasty (THA). The purpose of our study was to apply this Hip-Spine Classification to a sample of Australian patients undergoing THA surgery to determine the local acetabular component positioning requirements. Additionally, we propose a modified algorithm for adjusting cup anteversion requirements. 790 patients who underwent THA surgery between January 2021 and June 2022 were assessed for anterior pelvic plane tilt (APPt) and sacral slope (SS) in standing and relaxed seated positions and categorized according to their spinal stiffness and flatback deformity. Spinal stiffness was measured using pelvic mobility (PM); the ΔSS between standing and relaxed seated. Flatback deformity was defined by APPt <−13° in standing. As in Vigdorchik et al., PM of <10° was considered a stiff spine. For our algorithm, PM of <20° indicated the need for increased cup anteversion. Using this approach, patient-specific cup anteversion is increased by 1° for every degree the patient's PM is <20°. According to the Vigdorchik simple Hip-Spine classification groups, we found: 73% Group 1A, 19% Group 1B, 5% Group 2A, and 3% Group 2B. Therefore, under this classification, 27% of Australian THA patients would have an elevated risk of dislocation due to spinal deformity and/or stiffness. Under our modified definition, 52% patients would require increased cup anteversion to address spinal stiffness. The Hip-Spine Classification is a simple algorithm that has been shown to indicate to surgeons when adjustments to acetabular cup anteversion are required to account for spinal stiffness or flatback deformity. We investigated this algorithm in an Australian population of patients undergoing THA and propose a modified approach: increasing cup anteversion by 1° for every degree the patient's PM is <20°.
Over 1% of the global population suffers with ankle osteoarthritis (OA), yet there is limited knowledge on the changes to subchondral bone with OA. In other joints, it has been shown that bone becomes osteosclerotic, with fewer, thicker trabeculae that become hypomineralised, causing an increased apparent bone volume fraction (BV/TV). Microstructural alterations reduce overall joint strength, which may impact the success of late-stage surgical interventions, such as total ankle arthroplasty (TAA). Previous ankle studies have evaluated changes to cartilage, bone plate and bone morphology with OA, hence this study aimed to characterise changes to trabecular architecture. Three ankle joints were isolated from non-diseased cadaveric feet (three males: 43, 50 and 57 years, MEEC 18-027). Cylindrical subchondral bone specimens (N=6, 6.5 mm Ø) were extracted from the tibial plafond. Osteoarthritic bone samples (N=6, distal tibia) were sourced from local patients (three males: 65, 58 and 68 years, NREC 07/Q1205/27) undergoing TAA surgery. Specimens were imaged using µCT at a 16 µm isotropic resolution (µCT-100 ScanCo Medical). Virtual cores of bone (6.5 mm Ø) were extracted from the image data of the osteoarthritic specimens and trimmed to a height of 4 mm. BoneJ was used to evaluate key morphological indices: BV/TV; anisotropy (DA); trabecular thickness (Tb.Th); trabecular density (Conn.D) and ellipsoid factor (EF) which characterises rod/plate geometry. Differences between the two groups of specimens were evaluated using a t-test with Bonferroni correction.Abstract
Objectives
Methods
The current ‘gold’ standard surgical intervention for critical size bone defect repair involves autologous bone grafting, that risks inadequate graft containment and soft tissue invasion. Here, a new regenerative strategy was explored, that uses a barrier membrane to contain bone graft. The membrane is designed to prevent soft tissue ingrowth, whilst supporting periosteal regrowth, an important component to bone regeneration. This study shows the development of a collagen-based barrier membrane supportive of periosteal-derived mesenchymal stem cell (P-MSC) growth. P-MSC-homing barrier membranes were successfully obtained with nonaligned fibres, via free-surface electrospinning using type I collagen and poly(E-caprolactone) in 1,1,1,3,3,3-Hexafluoro-2-propanol. Introduction of collagen in the electrospinning mixture was correlated with decreased mean fibre diameter (d: 319 nm) and pore size (p: 0.2–0.6 μm), with respect to collagen-free membrane controls (d: 372 nm; p: 1–2 μm). Consequently, as the average MSC diameter is 20 μm, this provides convincing evidence of the creation of a MSC containment membrane. SEM-EDX confirmed Nitrogen and therefore collagen fibre localisation. Quantification of collagen content, using Picro Sirius Red dye, showed a 50% reduction after 24 hours (PBS, 37 °C), followed by a drop to 25% at week 3. The collagen-based membrane has a significantly higher elastic modulus compared to collagen-free control membranes. P-MSCs attached and proliferated when grown onto collagen-based membranes, imaged using confocal microscopy over 3 weeks. A modified transwell cell migration assay was developed, using MINUSHEET® tissue carriers to assess barrier functionality. In line with the matrix architecture, the collagen-based membrane proved to prevent cell migration (via confocal microscopy) in comparison to the migration facilitating positive control. The aforementioned results obtained at molecular, cellular and macroscopic scales, highlight the applicability of this barrier membrane in a new ‘hybrid graft’ regenerative approach for the surgical treatment of critical size bone defects.
Periprosthetic infections are leading causes of revision surgery resulting in significant increased patient comorbidities and costs. Considerable research has targeted development of biomaterials that may eliminate implant-related infections.1 This Several surface treated silicon nitride (Si3N4, Introduction
Materials and Methods
The exact mechanisms leading to tendinopathies and tendon ruptures remain poorly understood while their occurrence is clearly associated with exercise. Overloading is thought to be a major factor contributing to the development of tendon pathologies. However, as animal studies have shown, heavy loading alone won't cause tendinopathies. It has been speculated, that malfunctioning adaptation or healing processes might be involved, triggering tendon tissue degeneration. By analysing the expression of the entirety of degrading enzymes (degradome) in pathological and non-pathological, strained and non-strained tendon tissue, the aim of this study was to identify common or opposite patterns in gene regulation. This approach may generate new targets for future studies. RNA was extracted from different tendon tissues: normal (n=7), tendinopathic (n=4) and ruptured (n=4) Achilles tendon; normal (n=4) and tendinopathic (n=4) posterior tibialis tendon; normal hamstrings tendon with or without subjection to static strain (n=4). The RNA was reverse transcribed, then pooled per group The expression of 538 protease genes was analysed using Taqman low-density array quantitative RT-PCR. To be considered relevant, changes had to be at least 4fold and measurable at a level below 36 Cts.Introduction
Materials and Methods
Tendinopathies are debilitating and painful conditions. They are believed to result from repetitive overuse, which can create micro-damage that accumulates over time, and initiates a catabolic cell response. The aetiology of tendinopathy remains poorly understood, therefore the ideal treatment remains unclear. However, current data support the use of eccentric exercise as an effective treatment. In a previous study, we have shown that eccentric loading generates perturbations in the tendon at 10Hz, which is not present during other less effective loading regimes. Consequently, we hypothesis that 10Hz loading initiates an increased anabolic response in tenocytes, that can promote tendon repair. Human tenocytes from healthy hamstring tendons and tendinopathic Achilles tendons were derived by collagenase digest and outgrowth respectively. Tenocytes were seeded into 3D collagen gels. The gels were fixed in custom-made chambers and placed in an incubator for 24hrs whilst gene expression stabilised. After 24hrs, cyclic uniaxial strain at 1% ± 1% was applied to the cells, at either 1Hz (n=4) or 10Hz (n=4) using a Bose loading system. After 15 minutes of cyclic strain, the samples were maintained in chambers under 1% static strain for 24 hrs after which gene expression was characterised using RT-PCR.Introduction
Materials and Methods
We have shown that integrin mRNA expression is regulated by the application of mechanical load. This indicates that mechanical loading may modify cell sensitivity to perceive further load through increased interaction with the ECM. Tendinopathies are a range of diseases characterised by pain and insidious degeneration. Although poorly understood, onset is often associated with physical activity. We have previously investigated the regulation by mechanical strain of metalloproteinase gene expression in human tenocyte in a 3D collagen matrix. Integrins are important in cellular interaction with the ECM and are reported to mediate mechanotransduction in various non-tendon tissues. We have reported that TGFbeta activation is a key player in the regulation of metalloproteinases in response to mechanical load, which may be mediated by integrins. This project aims to investigate the effect of cyclic loading and TGFbeta stimulation on integrin expression by human tenocytes, in collagen and fibrin matrices.Summary Statement
Introduction
We hypothesise that the Masquelet induced membrane used for the reconstruction of large bone defects were likely to involve mesenchymal stem cells (MSCs), given the excellent resultant skeletal repair. This study represents the first characterisation in humans of the induced membrane formed as a result of the Masquelet technique. Induced membranes and matching periosteum were harvested from 7 patients. Cytokines (BMP2, VEGF, SDF1) and cell lineage markers (CD31, CD271, CD146) were studied by immunohistochemisty. Flow cytometry was used to measure the cellularity and cellular composition. MSCs were enumerated using a colony forming unit fibroblast assay. In expanded cultures, a 96-gene array card was used to assess their transcriptional profile. Alkaline phophatase, alizarin red and calcium assays were employed to measure their in vitro osteogenic potential Membrane was more cellular(p=0.028), had more MSC phenotype(p=0.043) compared to matched periosteum. The molecular profiles were similar, except for 2-fold abundance of SDF-1 in membrane (p=0.043)compared to periosteum. Membrane and periosteum had a similar proportion of endothelial cells and CFU-F colonies; expanded MSCs from both sources were highly osteogenic.Methods
Results
The concept of “bone graft expanders” has been popularised to increase the volume and biological activity of the implanted Material. Orthoss® granules support exogenously seeded MSCs and attract neighbouring host MSCs.Introduction
HYPOTHESIS
The aim of this study was to investigate PDGF release in the peripheral circulation following trauma and to correlate it with the numbers of MSCs in iliac crest bone marrow (BM) aspirate. Trauma patients with lower extremity fractures (n=18, age 21–64 years) were recruited prospectively. Peripheral blood was obtained on admission, and at 1, 3, 5 and 7 days following admission. The serum was collected and PDGF was measured using ELISA. Iliac crest (BM) aspirate (20ml) was obtained on days 0–9 following admission. MSCs were enumerated using standard colony-forming unit fibroblasts (CFU-F) assay.Objective
Methods
MSCs have long promised benefits of synthesising bone/cartilage, treating non-unions and potentially accelerating fracture repair. This potential has been tempered by MSC scarcity in the ‘gold-standard’ iliac crest bone marrow aspirate (ICBMA) and the resulting need to expand numbers via cell-culture. Culture of MSCs is time-consuming, expensive and results in cells with a reduced differentiation capacity. The reamer-irrigator-aspirator (RIA) is an innovation designed to reduce intra-medullary (IM) pressures during reaming of long-bones via continuous irrigation and suction. Aspirated contents are passed via a coarse filter, which traps bony-fragments before moving into a ‘waste’ bag - from which MSCs have been previously isolated. We examined liquid and solid phases found in this ‘waste’, performed a novel digestion of the solid phase and made a comparative assessment in terms of number, phenotype and differentiation capacity with matched ICBMA. The filtrate ‘waste’ bag from RIA reaming (6 patients) was filtered (70μm) and the solid fraction digested for 60min (37°C) with collagenase. MSCs were isolated from liquid & solid fractions and from 10ml matched ICBMA. Enumeration of MSCs was achieved via colony-forming-unit-fibroblast (CFUF) assay and flow-cytometry on fresh sample using CD45low, CD271+. MSCs were cultured by virtue of their plastic adherence and passaged in standard, non-haematopoietic media. Passage (P2) cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages with their phenotype assessed with flow cytometry CD33 CD34 CD45 CD73 CD90 CD105.Introduction
Methods
Although chondrocytes have been used for autologous implantation in defects of articular cartilage, limited availability and donor-site morbidity have led to the search for alternative cell sources. Mesenchymal stem cells from various sources represent one option. The infrapatellar fat-pad is a promising source. Advantages include low morbidity, ease of harvest and ex-vivo evidence of chondrogenesis. Expansion of MSCs from human fat-pad in FGF-2 has been shown to enhance chondrogenesis. To further elucidate this process, we assessed the role of TGF-?3, FGF-2 and oxygen tension on growth kinetics of these cells during expansion. Infrapatellar fatpads were obtained from 4 donors with osteoarthritis. Cells were expanded in various media formulations (STD, FGF, TGF and FGF/TGF) at both 20% and 5% oxygen tensions. Colony forming unit fibroblast assays were performed for each expansion group and assessed with crystal violet staining. Cell aggregates from each group underwent chondrogenic differentiation in 5% and atmospheric oxygen tension. Pellets were analyzed on day 21. 5% Oxygen tension during expansion increased the colony size for both FGF and FGF/TGF groups. Cells expanded in FGF/TGF proliferated more rapidly. Biochemical analysis revealed that cells expanded in FGF-2 had higher glycosaminoglycan synthesis rates, a marker for chondrogenesis. Differentiation at 5% pO2 led to higher levels of sGAG but its effect was generally less potent compared to expansion in FGF-2.Methods
Results
Iliac crest bone marrow aspirate (ICBMA) is frequently cited as the ‘gold-standard’ source of MSCs. Mesenchymal stem cells have been shown to reside within the intramedullary (IM) cavities of long-bones and a comparative assessment with ICBMA has not yet been performed. Aspiration of the IM cavities of 6 patients' femurs with matched ICBMA was performed. The long-bone-fatty-bone-marrow (LBFBM) aspirated was filtered (70μm) and the solid fraction digested for 60min (37°C) with collagenase. Enumeration was performed via the colony-forming-unit-fibroblast (CFU-F) assay and using the CD45low CD271+ phenotype via flow-cytometry. Passaged (P2) cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages with their phenotype assessed using flow-cytometry CD33 CD34 CD45 CD73 CD90 CD105.Introduction
Methods
Fracture healing represents a physiological process regulated by a variety of signalling molecules, growth factors and osteogenic progenitor cells. Bone healing following trauma is associated with increased serum concentrations of several pro-inflammatory and angiogenic growth factors1. Platelet-derived growth factor (PDGF) has been shown to stimulate mesenchymal stem cell (MSC) proliferation in vitro. However, the in vivo relationship between the levels of PDGF and the numbers of MSCs in humans has not yet been explored. The aim of this study was to investigate PDGF release in the peripheral circulation following trauma and to correlate it with the numbers of MSCs in iliac crest bone marrow (BM) aspirate and in peripheral blood. Trauma patients with lower extremity fractures (n=12, age 18-63 years) were recruited prospectively. Peripheral blood was obtained on admission, and at 1, 3, 5 and 7 days following admission. The serum was collected and PDGF was measured using the enzyme-linked immuno-sorbent assay (ELISA) technique. Iliac crest (BM) aspirate (20ml) and peripheral blood (PB) (20ml) was obtained on days 0-9 following admission. MSCs were enumerated using standard colony-forming unit fibroblasts (CFU-F) assay.Background and objectives
Methods
Iliac crest bone marrow aspirate (ICBMA) is frequently cited as the ‘gold-standard’ source of MSCs. MSCs have been shown to reside within the intramedullary (IM) cavities of long-bones [Nelea, 2005] however a comparative assessment with ICBMA has not yet been performed and the phenotype of the latter compartment MSCs remains undefined in their native environment. Aspiration of the IM cavities of 6 patients' femurs with matched ICBMA was performed. The long-bone-fatty-bone-marrow (LBFBM) was filtered (70μm) to separate liquid and solid fractions and the solid fraction was briefly (60min, 37oC) digested with collagenase. MSC enumeration was performed using the colony-forming-unit-fibroblast (CFU-F) assay and quantification of cells with the CD45low CD271+ phenotype by flow-cytometry. [Jones 2002, Buhring 2007] MSCs were cultured and standard expansion media and passage 2 cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages.Introduction
Methods
Therapeutic exploitation of MSCs in orthopaedics has been tempered by their scarcity within ‘gold-standard’ iliac crest bone marrow aspirate (ICBMA) and the resulting need to expand cells in vitro. This is time-consuming, expensive and results in cells with a reduced differentiation capacity. [Banfi 2000] The RIA is a device that provides continuous irrigation and suction during reaming of long bones. Aspirated contents pass via a filter, trapping bony-fragments, before moving into a ‘waste’ bag, from which MSCs have been previously isolated. [Porter 2009] We hypothesised that ‘waste’ RIA bag contains more MSCs than a standard aspirated volume of ICBMA (30 ml). We further hypothesised than a fatty solid phase within this ‘waste bag’ contains many MSCs trapped within the adipocyte-rich stromal network and hence requiring an enzymatic digestion for their efficient release [Jones 2006]. The discarded filtrate ‘waste’ bag that contained saline from marrow cavity irrigation procedure from RIA reaming (7 patients) was filtered (70μm) and the solid fraction digested for 60min (37oC) with collagenase. MSC enumeration was performed using the colony-forming-unit-fibroblast (CFU-F). Following culture in standard expansion media, passage 2 cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages and their phenotype was assessed using flow cytometry. ICBMA from the same patients was used as controls.Introduction
Methods
The scarcity of mesenchymal stem cells (MSCs) in iliac crest bone marrow aspirate (ICBMA), and the expense and time in culturing cells, has led to the search for alternative harvest sites. The reamer-irrigation-aspirator (RIA) provides continuous irrigation and suction during reaming of long bones. The aspirated contents pass via a filter, trapping bony fragments, before moving into a ‘waste’ bag from which MSCs have been previously isolated. We examined the liquid and solid phases, performed a novel digestion of the solid phase, and made a comparative assessment in terms of number, phenotype and differentiation capacity with matched ICBMA. The solid fraction from the filtrate was digested for 60 minutes at 37°C with collagenase. Enumeration was performed via the colony-forming unit fibroblast (CFU-F) assay. Passage (P2) cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages, and their phenotypes assessed using flow cytometry (CD33, CD34, CD45, CD73, CD90, and CD105). MSCs from the RIA phases were able to differentiate at least as well as those from ICBMA, and all fractions had phenotypes consistent with other established sources. The median number of colonies for the three groups was: ICBMA = 8.5 (2 to 86), RIA-liquid = 19.5 (4 to 90), RIA-solid = 109 (67 to 200) per 200 μl. The mean total yield of cells for the three groups was: ICBMA = 920 (0 to 4275), RIA-liquid = 114 983 (16 500 to 477 750), RIA-solid = 12 785 (7210 to 28 475). The RIA filtrate contains large numbers of MSCs that could potentially be extracted without enzymatic digestion and used for bone repair without prior cell expansion.
A combination of stem cell therapy and tissue engineering is emerging as one of the most promising approaches for skeletal tissue repair and regeneration. Osteoinduction of human bone marrow mesenchymal stem cells (MSCs) is initiated through local signals or growth factors, of which the bone morphogenetic proteins (BMPs) are the best characterised. Cytomodulin-1 (CM-1), a synthetic heptapeptide with functional similarity to members of the TGF-B super family, has been classified as a novel growth factor associated with osteoinduction of MSCs. However, the effects of CM-1 on human bone MSCs are still unclear. The aim of this study was to determine any effects for CM-1 and its scrambled control (CM-1 SCRAM) on the proliferation and differentiation of human bone marrow MSCs along the osteogenic lineage. Primary human bone marrow MSCs were cultured in the presence of CM-1 and CM-1 SCRAM at a range of concentrations (10-8M – 10-6M) in vitro for up to three weeks. 100 ng/mL of recombinant human BMP-2 (rhBMP-2) was used as a positive control. At the end of the culture period, histological and biochemical assays were carried out on the cultures. Biochemical assays revealed that 10-7M of CM-1 significantly stimulated alkaline phosphatase specific activity compared with the negative control group (P<
0.05) in a similar way to the rhBMP-2 positive control group. These data were supported by an observed increase in positive alkaline phosphatase staining in the 10-7M of CM-1 and rhBMP-2 treated cells. However, total DNA content was not significantly different between any of the groups. This study indicated the potential of using CM-1 as an osteogenic growth factor for skeletal tissue regeneration which may provide an alternative approach to meet the major clinical need in orthopaedics and craniofacial surgery. * Cytomodulin-1 and the scrambled control were genuine gifts from Professor (emeritus) Rajendra S. Bhatnagar at the Department of Bioengineering, University California Berkley, USA.
