RNA-Seq or whole transcriptome shotgun sequencing has been adopted in the last years as a reference technique to determine the presence and the quantity of different species of RNA in determined biological samples, thanks to it allows the identification every single RNA species transcribed from a reference genome. Meta-profiling takes advantage of the public availability of an increasing set of RNA-Seq data produced by different laboratories to summarize the expression levels of the different RNA species of many samples according to their biological context, giving the opportunity to perform comparisons on the gene expression profiles of different tissues by integrating data derived from a high number of studies. By using Genevestigator™; a platform which integrates RNA-Seq data into meta-profiles, we have performed a comparison between the gene expression profiles of bone, cartilage, muscle tendon and skin by means of interrogating its database with different gene sets and families with relevance to the function of the tissues of the musculoskeletal system. The collagen gene family and genes coding for proteoglycans, matrix metalloproteinases and tissue inhibitors of metalloproteinases, mechanotransduction-related proteins and signalling pathways involved in tissue development and differentiation have been analysed. Hierarchical clustering for every gene set was performed for the understanding the differences and similarities between the different tissues included in the analyses. The results of this study will help to improve our understanding of the musculoskeletal system, and will help to identify new biomarkers and signalling pathways of specific relevance for the bone, cartilage, muscle and tendon

Osteoarthritis is the most prevalent joint disease, causing severe pain, deformity and a loss of mobility. Low back pain (LBP), frequently associated with degeneration of the intervertebral disc (IVD), is the No.1 cause of Years Lived with Disability. Age is a major risk factor for both conditions. However, the reasons why susceptibility to these conditions increases with age are poorly understood. The circadian (24 hourly) clocks in the brain and periphery direct key aspects of physiology through rhythmic control of tissue-specific sets of downstream genes. Work from our group focuses on the roles of circadian clocks in the articular cartilage and IVD. We show that the daily rhythm in these tissues becomes dampened and out-of-phase during ageing. Further, our data identify circadian clock disruption in cartilage and IVD as a new target of inflammation. Moreover, we show that mice with targeted knockout of an essential clock gene (BMAL1) in chondrocytes and disc cells have profound, yet tissue-specific degeneration in the articular cartilage and IVD. These findings implicate the local skeletal clock as a key regulatory mechanism for tissue homeostasis. This new avenue of research holds potential to better understand, and eventually treat these debilitating conditions.

Imaging can provide valuable information about the function of tissues and organs. The capacity for detecting and measuring imaging biomarkers of biological activities, allows for a better understanding of the pathophysiology of any process in the human body, including the musculoskeletal system. This is of particular importance in oncologic, metabolic and rheumatologic diseases, but not limited to these. In the domain of the musculoskeletal system, functional imaging also means to be able to address biomechanical evaluations. Weight-bearing imaging and dynamic studies have a prominent role. All imaging techniques (X-rays, CT, MR, ultrasound) are in demand, and offer different applications, specific equipment and novel methods for addressing this. Functional imaging is also essential to drive minimally invasive treatments – i.e. interventional radiology, and new treatment approaches move together with the advances on imaging guidance methods. On both the diagnostic and the interventional side, the increasing availability of dedicated equipment and the development of specific imaging methods and protocols greatly helps the transition from research to clinical practice

Our musculoskeletal system has a limited capacity for repair. This has led to increased interest in the development of tissue engineering and biofabrication strategies for the regeneration of musculoskeletal tissues such as bone, ligament, tendon, meniscus and articular cartilage. This talk will demonstrate how different musculoskeletal tissues, specifically cartilage, bone and osteochondral defects, can be repaired using emerging biofabrication and 3D bioprinting strategies. This will include examples from our lab where cells and/or growth factors are bioprinted into constructs that can be implanted directly into the body, to approaches where biomimetic tissues are first engineered in vitro before in vivo implantation. The efficacy of these different biofabrication strategies in different preclinical studies will be reviewed, and lessons from the relative successes and failures of these approaches to tissue regeneration will be discussed

Osteotomies in the musculoskeletal system are joint preserving procedures to correct the alignment of the patient. In the lower limb, most of the pre-operative planning is performed on full leg weightbearing radiographs. However, these images contain a 2-dimensional projection of a 3-dimensional deformity, lack a clear visualization of the joint surface and are prone to rotational errors during patient positioning. Weightbearing CT imaging has demonstrated to overcome these shortcomings during the first applications of this device at level of the foot and ankle. Recent advances allow to scan the entire lower limb and novel applications at the level of the knee and hip are on the rise. Here, we will demonstrated the current techniques and 3-dimensional measurements used in supra- and inframalleolar osteotomies around the ankle. Several of these techniques will be transposed to other parts in the lower limb to spark future studies in this field

Geometric deep learning is a relatively new field that combines the principles of deep learning with techniques from geometry and topology to analyze data with complex structures, such as graphs and manifolds. In orthopedic research, geometric deep learning has been applied to a variety of tasks, including the analysis of imaging data to detect and classify abnormalities, the prediction of patient outcomes following surgical interventions, and the identification of risk factors for degenerative joint disease. This review aims to summarize the current state of the field and highlight the key findings and applications of geometric deep learning in orthopedic research. The review also discusses the potential benefits and limitations of these approaches and identifies areas for future research. Overall, the use of geometric deep learning in orthopedic research has the potential to greatly advance our understanding of the musculoskeletal system and improve patient care

Non-linear methods in statistical shape analysis have become increasingly important in orthopedic research as they allow for more accurate and robust analysis of complex shape data such as articulated joints, bony defects and cartilage loss. These methods involve the use of non-linear transformations to describe shapes, rather than the traditional linear approaches, and have been shown to improve the precision and sensitivity of shape analysis in a variety of applications. In orthopedic research, non-linear methods have been used to study a range of topics, including the analysis of bone shape and structure in relation to osteoarthritis, the assessment of joint deformities and their impact on joint function, and the prediction of patient outcomes following surgical interventions. Overall, the use of non-linear methods in statistical shape analysis has the potential to advance our understanding of the relationship between shape and function in the musculoskeletal system and improve the diagnosis and treatment of orthopedic conditions

In the context of regenerative medicine for the treatment of musculoskeletal pathologies mesenchymal stromal cells (MSCs) have shown good results thanks to secretion of therapeutic factors, both free and conveyed within the extracellular vesicles (EV), which in their totality constitute the “secretome”. The portfolio and biological activity of these molecules can be modulated by both in vitro and in vivo conditions, thus making the analysis of these activities very complex. A deep knowledge of the targets regulated by the secretome has become a matter of fundamental importance and a homogeneous and complete molecular characterization is still lacking in the field of applications for the musculoskeletal system. Therefore, the aim of this work was to characterize the secretome obtained from adipose-derived MSCs (ASCs), and its modulation after pre-conditioning of the ASCs. Pre-conditioning was done by culturing cells in the presence of i) high levels of IFNγ, as proposed for the production of clinical grade secretome with enhanced regenerative potential, ii) low levels of inflammatory stimuli, mimicking conditions found in the osteoarthritis (OA) synovial fluid. Furthermore, EVs ability to migrate within cartilage, chondrocyte and synoviocytes obtained from OA patients was evaluated. The data showed that more than 50 cytokines / chemokines and more than 200 EV-microRNAs are detectable at various intensity levels in ASCs secretomes. The majority of the most abundantly present molecules are involved in the remodelling of the extracellular matrix and in the homeostasis and chemotaxis of inflammatory cells including macrophages, which in OA are often characterized by an M1 inflammatory polarization, promoting their transition to an M2 anti-inflammatory phenotype. Inflammatory priming with IFNγ and synovial fluid-like conditions were able to further increase the ability of the secretome to interact with inflammatory cells and modulate their migration. Finally, the penetration of the EVs in the cartilage explants resulted a rapid process, which begins a few minutes after administration of the EVs that are able to reach a depth of 30-40 μm in 5 hours. The same capacity for interaction was also verified in chondrocytes and synoviocytes isolated from the cartilage and synovial membrane of OA patients. Thanks to the soluble factors and EV-microRNAs, the ASCs secretome has shown a strong propensity to modulate the inflammatory and degenerative processes that characterize OA. The inflammatory pre-conditioning through high concentrations of inflammatory molecules or in conditions similar to the synovial fluid of OA patients was able to increase this capacity by increasing their chemotactic power. The microscopy data also support the hypothesis of the ability of MSC-EVs to influence the chondrocytes residing in the ECM of the cartilage and the synovial cells of the synovial membrane through active interaction and the release of their therapeutic content

The human musculoskeletal system is a biological composite of hard and soft material phases organized into a complex 3D structure. The replication of mechanical properties in 3-dimensional space, so called ‘4D’ techniques, therefore promises next-generation of prosthetics and engineering structures for the musculoskeletal system. Approaches using in situ indentation of tissue correlated with micro computed tomography (μCT) are used here to provide a 4D data set that is representative of the native tissue at high fidelity. Multi-material 3D printing is exploited to realize the collected 4D data set by using materials with a wide range of mechanical properties and printing structures representative of native tissue. We demonstrate this correlative approach to reproduce bone structures and highlight a workflow approach of indentation, μCT and 3D printing to potentially mimic any structure found in the musculoskeletal system. Structures in the human musculoskeletal system, such as bone [1] and tendon-bone connective tissue [2], can be considered as complex composites of hard and soft materials. Development of prosthetics capable of replacing body parts lost to trauma, disease or congenital conditions requires the accurate replication of the required body part. 3D printing promises considerable advantages over other manufacturing methods in mimicking native tissue, including the ability to produce complex structures [3]. However, accurate representation of whole body parts down to tissue microstructures requires correlative approaches where mechanical properties in 3-dimensional space are known. The objective of this study is to apply in situ indentation, correlate to 3D imaging of bone using μCT and finally 3D print mimicked structures. Samples of bovine compact bone were imaged at high resolution using μCT (Xradia Versa 510, Zeiss, USA). A custom build in situ micro indentation setup within the μCT was used to map the mechanical properties of the bone at multiple positions. Correlation between sample x-ray attenuation and corresponding elastic modulus found from indentation was established. Data was converted to a 4D data set of elastic modulus values in 3D space, segmented and exported to the 3D printer. An inkjet 3D printer (Projet 5500X, 3D Systems, USA) was used to print materials with a range of mechanical properties that approach those found in the native bone material. Macroscopic testing on both bone samples and 3D printed samples were carried out using standard compression (Instron, UK). Preliminary results indicated similarity between 3D printed structures and native bone tissue. Macroscopic testing of bone samples and 3D printed equivalents showed additional similarities in stress-strain behaviour. Our preliminary work presented here indicates that the workflow of 3D imaging correlated to point mechanical measurements using indentation is suitable to give a 4D dataset that is representative of the native bone tissue. 3D printing is able to produce structures that start to mimick bone but are critically dependent on the data segmentation, particularly averaging imaging data to a resolution that is appropriate for the 3D printer

Calcium is an important element for a wide range of physiological functions including muscle contraction, neuronal activity, exocytosis, blood coagulation and cell communication. In the musculoskeletal system calcium is crucial for the structural integrity of bones, teeth, intervertebral disc and articular cartilage. At the cellular level calcium acts as a second messenger. Calcium signalling uses intracellular calcium ions to drive intracellular communication and signal transduction processes. When calcium enters the cell it exerts allosteric regulatory effects on many enzymes and proteins. Examining the role of calcium in chondrocyte biology is important for understanding the role for this divalent ion in the metabolic modulation of chondrocyte function in health and disease. This includes the study of calcium transport systems such as channels, transporters and pumps involved in calcium homeostasis in chondrocytes and how existing pharmacological drugs act on these transport systems. L-type calcium channel blockers are drugs used as cardiac antiarrhythmics or antihypertensives, depending on whether the drugs have higher affinity for the heart (the phenylalkylamines, like verapamil), or for the blood vessels (the dihydropyridines, like nifedipine). L-type calcium channels are present in many musculoskeletal tissues including skeletal muscle, smooth muscle, bone and cartilage. L-type calcium channel inhibitors like nifedipine used for the treatment of some forms of hypertension modulate calcium-mediated events in chondrocytes under dynamic loading, thus affecting metabolism, osmotic responses and extracellular matrix turnover in cartilage. The aim of our work is to understand the impact of L-type calcium channel inhibitors used for the treatment of hypertension on chondrocytes and on the chondrogenic differentiation of bone marrow derived mesenchymal stem cells (MSCs). This knowledge will enhance our understanding of the development of osteoarthritis (OA) and may lead to new opportunities for chondroprotection and regenerative medicine for OA. We have used electrophysiology to demonstrate L-type calcium currents in chondrocytes immediately after pharmacological activation with the calcium channel opener Bay-K8644. We have also used immunohistochemistry to demonstrate expression of the a1C subunit Ca. v. 1.2 (CACNA1C) in human chondrocytes and MSCs. Inhibitors of L-type calcium channels such as nifedipine downregulate mitochondrial respiration and ATP production in MSCs but not in chondrocytes. Nifedipine inhibits proliferation of chondrocytes and enhances glycolytic capacity in chondrocytes, promoting glycolytic reserve in both MSCs and chondrocytes. Nifedipine can also stimulate chondrogenic differentiation in MSCs (with or without growth factors). Metabolic responses to nifedipine differs in mesenchymal stem cells and chondrocytes highlighting important metabolic differences between these cells. In summary, antihypertensive drugs such as nifedipine can affect the biological function of chondrocytes and MSCs and may modulate the course of OA progression and impact on cartilage repair

Mesenchymal Stromal Cells (MSC) have been proposed as a potential therapy for a broad range of diseases including those affecting the musculoskeletal system. MSCs have received market authorization for treatment of graft versus host disease and fistulizing Crohn's disease. In addition, there are clinical trials underway for diseases affecting all organ systems. GMP manufactured cells are required for these clinical trials and suitable facilities with regulatory approval are thus crucial for the translational process. In this presentation I will review the process whereby such a facility has been constructed at NUI Galway and discuss challenges in operations and sustainability. Researchers at REMEDI and spin out company Orbsen Therapeutics are currently involved in 7 clinical trials using MSCs, 4 of which are EU wide consortia funded by the EU Commission. The presentation will also discuss issues such as source of MSCs, cell sorting, use of bioreactors and xeno-free processes

Our musculoskeletal system has a limited capacity for repair. This has led to increased interest in the development of tissue engineering strategies for the regeneration of musculoskeletal tissues such as bone, ligament, tendon, meniscus and articular cartilage. This talk will review our attempts to use biomaterials and mesenchymal stem cells (MSCs) to bioprint functional articular cartilage and bone grafts for use in bone and joint regeneration. It will begin by describing how 3D bioprinting can be used to engineer biological implants mimicking the shape of specific bones, and how these bioprinted tissues mature into functional bone organs upon implantation into the body. Next, it will be demonstrated that different musculoskeletal injuries can be regenerated using 3D bioprinted implants, including large bone defects and osteochondral defects. The talk will conclude by describing how we can integrate biomaterials and MSCs into 3D bioprinting systems to engineer scaled-up tissues that could potentially be used regenerate entire diseased joints

The establishment of a proper musculoskeletal system depends on the well-organized and synchronized development of muscle, tendon and cartilage/bone. In tendon biology, a great progress in identifying tendon-specific genes (Scleraxis, Mohawk, Tenomodulin) had been made in the last decade. However, there are many open questions regarding the exact function of genes in tendon development and homeostasis. The purpose of this study was to perform a systematic review of publications describing tendon-related genes, which were studied in-depth and characterized by using knockout technologies and the respectively generated transgenic mouse. Method: Literature search was carried out in Pubmed using “tendon” and “mouse knockout” and “phenotype” and was not limited to year. Results: We report in a tabular manner, that from a total of 25 tendon-related genes, in 23 of the respective knockout mouse models phenotypic changes were detected. Additionally, in some of the models it was described at which developmental stages these changes appeared and progressed. Interestingly, so far only loss of Scleraxis and TGFbeta signaling led to severe tendon developmental phenotypes, while mice deficient for various proteoglycans, Mohawk, EGR1 and 2, and Tenomodulin exhibited mild phenotypes. This suggests that in general the tendon developmental program is well backup and specifically that among the members of the proteoglycan family there are clear compensatory effects. In future, it will be of great importance to discover additional master tendon transcription factors as well as genes that play indispensable roles in tendon development

Digital image correlation (DIC) is rapidly increasing in popularity in biomechanical studies of the musculoskeletal system. DIC allows the re-construction of full field displacement and/or strain maps of the surface of an object. DIC systems typically consist of two cameras focussing on the same region of interest. This constrains the angle between the cameras to be relatively narrow when studying specimens characterised by complex geometrical features, giving rise to concerns on the accuracy of the out of plane estimates of movement. The aim of this research was to compare the movement profiles of bony segments measured by DIC and by an optoelectronic motion capture system. Five porcine cervical spine segments (C2-C6) were obtained from the local butcher. These were stripped of all anterior soft tissues while the posterior structures were left intact. A speckle pattern was applied to the anterior aspect of the specimens, while custom made infrared clusters were rigidly attached to the 3 middle vertebral bodies (C3-C5). The specimens were mounted in a custom made impact rig which fully constrained C6 but allowed C2 to translate in the axial direction of the segment. Images were acquired at 4kHz, both for the DIC (Photron Europe Ltd, UK) and motion capture cameras (Qualisys Oqus 400, Sweden). The in-plane and out of plane displacements of each of the VBs were plotted as a function of time and the similarity between the curves thus obtained was analysed using the SPM1D technique which allowed a comparison to be made in terms of t-statistics. No statistical differences were found between the two techniques in all axis of movement, however the out of plane movements were characterised by higher variance which is attributed to the uncertainty arising from the near parallel positioning of the cameras in the experimental set-up

Tendons are dense connective tissues and critical components of the musculoskeletal system with known long repair process. Tissue engineering is a promising approach for achieving complete recovery of ruptured tendons. The most studies have focused on the combination of cells with various carriers; however, frequent times the biomaterials do not match the tissue organization and strength. For this reason, we first reviewed the literature for an alternative scaffold-free strategy for tendon engineering and second, we compared the cell sheet formation of two different cell types: bone marrow-derived mesenchymal stem cells (BM-MSCs) and tendon stem/progenitor cells (TSPCs). Methods: Literature search was performed in Pubmed using “tendon tissue engineering” and “scaffold-free” keywords and was limited to the last ten years. By using a 3-step protocol, BM-MSCs and TSPCs were induced to form cell sheets in 5 weeks. The sheets were compared by analysis for weight, diameter, cell density, tissue morphology (H&E and scoring) and cartilaginous matrix (DMMB and S.O. staining). Results: Scaffold-free models (cell sheets and pellet cultures) are available; however, further optimization is needed. Comparison between the two cell types clearly demonstrated that TSPCs form more mature cell sheet, while BMSCs form larger but less organized and differentiated sheet as judged by higher cell density and lower scoring outcome. Future efforts will focus on identifying mechanisms to speed BM-MSC sheet formation and maturation, which can in turn lead to development of new methodology for scaffold-free tendon tissue engineering

Entheses are the anchorage sites of tendons to bones in the musculoskeletal system. They have a unique microanatomy that allow smooth transfer of mechanical load through tendon to bone. However, entheses are prone to injury due to their small surface area. 1,2. The overall success rate of the current gold standard treatment (directly attaching the tendon to bone) is small. 3,4. Consequently, the aim of this study was to evaluate different hydrogels and their suitability for developing an in-vitro co-culture system to manufacture 3D tissue interfaces. To create a 3D in-vitro tissue interface, half-well plugs were created by pouring silicone in wells of a 24-well plate. When set, it was cut into halves to be used as half-well plugs, blocking one side of a culture well. A tendon-cell-encapsulated hydrogel was poured into the exposed half and, when set, the plug was removed and a bone-cell-encapsulated gel was added. Cells were fluorescently labelled to enable identification of cell types under fluorescent microscopy (Tendon – green, bone – red). The suitability of different hydrogels to form an in vitro tissue interface was evaluated: fibrin, agarose and gellan. This study demonstrates that 3D co-cultures can be manufactured in-vitro. The novel system enabled the culture of two cell types (bone/tendon) in direct contact, creating an in-vitro interface. In addition, this study shows that fibrin gel supports cell morphology, while both cell types failed to show normal morphology in agarose and gellan. Further studies evaluating cell viability in these hydrogels are currently underway

Osteoporosis is an international health and financial burden of ever increasing proportions. Current treatments limit the rate of bone resorption and reduce fracture risk, however they are often associated with significant and debilitating side effects. The most commonly used therapies also do not stimulate osteoblast activity. Much current research focus is aimed at the metabolic and epigenetic pathways involved in osteoporosis. MicroRNAs have been shown to play an important role in bone homeostasis and pathophysiological conditions of the musculoskeletal system. Upregulation of specific microRNAs has been identified in-vivo in osteoporotic patients. It is hypothesized that modulation of specific mircoRNA expression may have a key role in future targeted therapies of musculoskeletal diseases. The assessment and analysis of their potential therapeutic use in Osteoporosis is of great importance, due to the burden of the disease. We have developed a 3D osteoporotic model from human bone marrow, without the use of scaffold. Magnetic nanoparticles are utilised to form spheroids, which provides a closer representation of the in-vivo environment than monolayer culture. This model will provide the basis for analysing future microRNA experiments to assess the potential upregulation of osteoblastogenesis without cessation of osteoclast activity. The results of initial monolayer and spheroid experiments will be presented. Optimisation of the osteoporotic bone marrow culture conditions, involving response to differentiation medias, analysis of adipose and bone markers and cell migration in spheroid culture will be displayed. Quantitative and qualitative results, including fluorescence microscopy and in cell western, assessing the monolayer and spheroid cultures will be presented. The development of a pseudo osteoporosis model from healthy bone marrow will also be discussed. This model will form a basis of future work on miRNA targeting

Osteoporosis is an international health and financial burden of ever increasing proportions. Current treatments limit the rate of bone resorption and reduce fracture risk, however they are often associated with significant and debilitating side effects. The most commonly used therapies also do not stimulate osteoblast activity . 1,2,3. Much current research focus is aimed at the metabolic and epigenetic pathways involved in osteoporosis. MicroRNAs have been shown to play an important role in bone homeostasis and pathophysiological conditions of the musculoskeletal system. Up-regulation of specific microRNAs has been identified in-vivo in osteoporotic patients . 4,5. It is hypothesized that modulation of specific microRNA expression may have a key role in future targeted therapies of musculoskeletal diseases. The assessment and analysis of their potential therapeutic use in Osteoporosis is of great importance, due to the burden of the disease. We have developed a 3D osteoporotic model from human bone marrow, without the use of scaffold. Magnetic nanoparticles are utilised to form spheroids, which provides a closer representation of the in-vivo environment than monolayer culture. This model will provide the basis for analysing future microRNA experiments to assess the potential up-regulation of osteoblastogenesis without cessation of osteoclast activity. The results of initial monolayer and spheroid experiments will be presented. Optimisation of the osteoporotic bone marrow culture conditions, involving response to differentiation medias, analysis of adipose and bone markers and cell migration in spheroid culture will be displayed. Quantitative and qualitative results, including fluorescence microscopy and in cell western, assessing the monolayer and spheroid cultures will be presented. The development of a pseudo osteoporosis model from healthy bone marrow will also be discussed. This model will form a basis of future work on microRNA targeting. The development of improved therapies for osteoporosis is of great significance due to the predicted rise in incidence of the disease and associated fragility fractures. Targeted therapies, such as the manipulation of microRNA expression, offer the opportunity to increase osteoblastogenesis and decrease osteoclastogenesis, potentially without the associated side effects of older, systemic therapies. We believe our 3D human bone marrow derived osteoporotic model offers the closest relation to the in-vivo environment for assessment and manipulation of microRNA expression

Introduction. Tendons and ligaments (TLs) play key roles in the musculoskeletal system. However, they are commonly damaged due to age-related wear and tear or torn in traumatic/sport related incidents resulting in pain and immobility. TLs contain cells and extracellular matrix (ECM) comprised of collagen, elastin, glycoproteins and proteoglycans. Although TLs are composed of similar components, their precise composition and arrangement of matrix macromolecules differ to provide specific mechanical properties and functions. To date little is known about how the main ECM proteins are arranged between the two tissue types. This data will provide essential information on fundamental structure of TLs leading to increased understanding of the function relationship between these tissue types. The aim of this study was to compare tendon-ligament differences in their ECM distribution of collagens, proteoglycans and elastic fibres. Materials and Methods. Anterior cruciate ligament (ACL) and long digital extensor tendon (LDET) were harvested from disease free cadaveric canine knee joints (n=3). Distributions of the main ECM components were assessed on longitudinal sections of ACL and LDET mid-substance. Antibody staining were assessed for collagen type I, III, VI, agreccan, versican, decorin, biglycan, elastin, fibrillin 1 and fibrillin 2. Results. Marked staining of collagen type I was present at fascicular regions, but also present at the interfascicular matrix (IFM). Collagen type III was present at the IFM of tendon, whilst in the ligament it was more widespread being located at both fascicles and IFM. In both TLs, collagen type VI was localised at IFM, but also present surrounding TL fibroblasts. A marked staining of aggrecan and versican was observed in ligament IFM, with pericellular staining of aggrecan present only in ligament. Decorin was found in both fascicular and IFM, whilst biglycan was occasionally present pericelullarly and at IFM in tendon. A similar pattern of elastic fibre distribution was found in both TLs. Discussion. This study has revealed a different ECM distribution of collagen type III, aggrecan, versican in ligament than when compared to tendon. These finding may relate to different functions between TLs and indicate that ligament is subjected to more compressive forces, resulting in different macromolecular arrangements that protect the tissue from damage

Finite element models of the musculoskeletal system have the possibility of describing the in vivo situation to a greater extent than a single in vitro experimental study ever could. However these models and the assumptions made must be validated before they can be considered truly useful. The object of this study was to validate, using digital image correlation (DIC) and strain gauging, a novel free boundary condition finite element model of the femur. The femur was treated as a complete musculoskeletal construct without specific fixed restraint acting on the bone. Spring elements with defined force-displacement relationships were used to characterize all muscles and ligaments crossing the hip and knee joints. This model was subjected to a loading condition representing single leg stance. From the developed model muscle, ligament and joint reaction forces were extracted as well as displacement and strain plots. The muscles with the most influence were selected to be represented in the simplified experimental setup. To validate the finite element model a balanced in vitro experimental set up was designed. The femur was loaded proximally through a construct representative of the pelvis and balanced distally on a construct representing the tibio-femoral joint. Muscles were represented using a cabling system with glued attachments. Strains were recorded using DIC and strain gauging. DIC is an image analysis technique that enables non-contact measurement of strains across surfaces. The resulting strain distributions were compared to the finite element model. The finite element model produced hip and knee joint reaction forces comparable to in vivo data from instrumented implants. The experimental models produced strain data from both DIC and strain gauging; these were in good agreement with the finite element models. The DIC process was also shown to be a viable method for measuring strain on the surface of the specimen. In conclusion a novel approach to finite element modeling of the femur was validated, allowing greater confidence for the model to be further developed and used in clinical settings