Introduction. Chondrocytes are enveloped within the pericellular matrix (PCM), a structurally intricate network primarily demarcated by the presence of collagen type VI microfibrils and perlecan, resembling a protective cocoon. The PCM serves pivotal functions in facilitating cell mechanoprotection and mechanotransduction. The progression of osteoarthritis (OA) is associated with alterations in the spatial arrangement of chondrocytes, transitioning from single strings to double strings, small clusters, and eventually coalescing into large clusters in advanced OA stages. Changes in cellular patters coincide with structural degradation of the PCM and loss of biomechanical properties. Here, we systematically studied matrix metalloproteinases (MMPs), their distribution, activity, and involvement in PCM destruction, utilizing chondrocyte arrangement as an OA biomarker. Methods. Cartilage specimens were obtained from 149 osteoarthritis (OA) patients, and selected based on the predominant spatial pattern of chondrocytes. Immunoassays were employed to screen for the presence of various MMPs (-1, -2, -3, -7, -8, -9, -10, -12, -13). Subsequently, the presence and activity of elevated MMPs were further investigated through immunolabeling, western blots and zymograms. Enzymatic assays were utilized to demonstrate the direct involvement of the targeted MMPs in the PCM destruction. Results. Screening revealed increased levels of MMP-1, -2, -3, -7, and -13, with their expression profile demonstrating a distinct dependency on the stage of degeneration. We found that MMP-2 and -3 can directly compromise the integrity of collagen type VI, whereas MMP-3 and MMP-7 disrupt perlecan. Conclusions. Presence of both pro- and active forms of MMP-2, -3, and -7 in OA-induced patterns, along with their direct involvement in collagen type VI and perlecan degradation, underscores their crucial role in early PCM destruction. Given the early stages of the disease already exhibit heightened MMP expression, this understanding could inform early targeted therapies aimed at arresting abnormal PCM remodelling. Acknowledgments. Faculty of Medicine of the University of Tübingen (grant: 2650-0-0)

Introduction. Osteoarthritis (OA) is a prevalent joint disorder characterized by cartilage degeneration, inflammation, and pain. Current treatments provide only symptomatic relief, necessitating novel molecular targets. The caspase family, known for its roles in apoptosis and inflammation regulation, may additionally influence crucial processes for cartilage homeostasis such as differentiation and proliferation. However, the specific roles of individual caspases in OA pathogenesis remain unclear. This study aims to investigate the involvement of the caspase family in OA and as potential targets for therapy, with a focus on caspase-1 and -8. Method. Chondrocytes from both healthy and OA donors were cultured in 2D and 3D culture models and stimulated with TNF-α or IL-1β. The expression and activation of caspase-1 and -8 was assessed using RT-PCR, ELISA. Transcriptome analysis of OA and healthy cartilage samples, along with Mendelian randomization (MR) analysis were conducted to explore the involvement of caspase family in OA and to assess its potential as therapeutic targets. Result. Higher expression levels of caspase-1, -8 were observed in OA cartilage compared to healthy cartilage. TNF-α stimulation increased their expression in both healthy and OA chondrocytes, while IL-1β had limited impact. Caspase-8 expression was causally associated with knee OA in MR analysis, suggesting a potential therapeutic target. The caspase-1 inhibitor VX-765 mildly reduced chondrocyte viability, with no significant effect in the presence of TNF-α. While the caspase-8 inhibitor Z-IETD-FMK exhibited slight enhancements in cell viability, these improvements were not statistically significant. Nevertheless, its effectiveness significantly increased in the presence of TNF-α. Conclusion. This study highlights the involvement of caspase-1 and caspase-8 in OA pathology, with caspase-8 emerging as a potential therapeutic target for knee OA treatment. Further investigation into the roles of caspase-1 and -8 in OA pathophysiology, including the efficacy and potential side effects of their corresponding inhibitors, is warranted. Acknowledgements. Funding Inter-Action/Inter-Excellence project (BTHA-JC-2022-36/LUABA22019)

Mesoporous bioactive glasses (MBGs) have been widely studied as bone regeneration systems, due to their bioactivity and ability to store and release therapeutic agents with specific biological functions. The incorporation of these nanomaterials into a thermosensitive hydrogel (TSH), in which a solution undergoes a sol-gel transition under physiological conditions, represents a promising approach to design multifunctional devices able to deliver selected molecules to pathological sites. In fact, this system can perfectly fit the defect cavity shape prior to the complete gelation, and acts as a carrier for therapeutic agents prolonged release in situ. This challenging concept is the underlying idea of the MOZART project, whose objective was to develop a library of MBGs containing different therapeutic ions and drugs, to be used as a new, smart platform technology for highly targeted therapies to enhance bone healing. The aim of this work is to investigate the bone regeneration potential of MBGs containing strontium ions (pro-osteogenic) and incorporated into thermosensitive poly(etherurethane)(PEU) based on Poloxamer407. In order to further increase the pro-osteogenic response, MBGs were also loaded with N-acetylcysteine (NAC). MBGs containing 2%mol of Sr. 2+. were prepared by an aerosol-assisted spray-drying method and NAC was loaded post-synthesis via an incipient wetness method. The PEU hydrogel (SHP407) was synthesized via a two-step procedure in nitrogen atmosphere. Particles were characterized (FE-SEM, N. 2. adsorption-desorption analysis, TGA, DSC, FT-IR and XRD) and then incorporated into the hydrogel. The hybrid systems rheological properties and stability in aqueous environment at 37°C, and its ability to co-release Sr. 2+. and NAC were analysed. After preliminary biological in vitro tests, a proof-of-concept rodent study was run to assess the ability of the resulting formulation as bone healing device. X-ray at 2 and 4-weeks post-surgery and µCT-analysis were used to evaluate the healing results in a rat osteotomy model of biologically impaired healing. Then, bones were processed for histological evaluation. Preliminary in vivo results demonstrated that incorporation of MBGs into a TSH is a promising strategy to design a multifunctional injectable formulation for in situ and sustained delivery of pro-osteogenic species enhancing bone regeneration

Infected non-unions of proximal femoral fractures are difficult to treat. If debridement and revision fixation is unsuccessful, staged revision arthroplasty may be required. Non-viable tissue must be resected, coupled with the introduction of an antibiotic-eluting temporary spacer prior to definitive reconstruction. Definitive tissue microbiological diagnosis and targeted antibiotic therapy are required. In cases of significant proximal femoral bone loss, spacing options are limited. We present a case of a bisphosphonate-induced subtrochanteric fracture that progressed to infected non-union. Despite multiple washouts and two revision fixations, the infection remained active with an unfavourable antibiogram. The patient required staged revision arthroplasty including a proximal femoral resection. To enable better function by maintaining leg length and offset, a custom-made antibiotic-eluting articulating temporary spacer, the Cement-a-TAN, was fabricated. Using a trochanteric entry cephalocondylar nail as a scaffold, bone cement was moulded in order to fashion an anatomical, patient-specific, proximal femoral spacer. Following resolution of the infection, the Cement-a-TAN was removed and a proximal femoral arthroplasty was successfully performed. Cement-a-TAN is an excellent temporary spacing technique in staged proximal femoral replacement for infected non-union of the proximal femur where there has been significant bone loss. It preserves mobility and maintains leg length, offset and periarticular soft-tissue tension

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

Multiple myeloma (MM) is a chronic, malignant B-cell disorder, with a less than 50% 5-year survival rate [1]. This disease is responsible for vertebral compression fractures (VCFs) in 34 to 64% of diagnosed patients [1], and at least 80% of MM patients experience pathological fractures [3]. Even though reduced DXA-derived bone mineral density (BMD) has been observed in MM patients with vertebral fractures [4], the current quantitative standard method is insufficient in MM due to the osteo-destructive bone changes. Finite-element (FE) analysis is a computational and non-destructive modeling and testing approach to determine bone strength using 3D bone models from CT images. Thus, this study aimed to assess the differences in FE-predicted critical fracture load in MM patients with and without VCFs in the thoracic and lumbar segments of the spine. Multi-detector CT (MDCT) images of two radiologically assessed MM patients (1 with VCFs and 1 without VCFs) were used to generate three-dimensional (3D) models of the whole spine. For each subject, the thoracic segments, 1 to 12 (T1-T12) and lumbar segments, 1 to 5 (L1-L5) were segmented and meshed. Heterogeneous, non-linear anisotropic material properties were applied by discretizing each vertebral segment into 10 distinct sets of materials. A compressive load was simulated by constraining the surface nodes on the inferior endplate in all directions, and a displacement load was applied on the surface nods on the superior endplate [2]. This analysis was performed using ABAQUS version 6.10 (Hibbitt, Karlsson, and Sorensen, Inc., Pawtucket, RI, USA). The MM subject with VCFs had originally experienced fractures in the T4, T5, T12, L1, and L5 segments whereas the MM subject without VCFs experienced none. The former displayed large and abrupt differences in fracture loads between adjacent vertebrae segments, unlike the latter, which exhibited progressive differences instead (no abrupt changes between adjacent vertebrae segments observed). Results from this preliminary study suggest that segments at high risk of fracture are collectively involved in an unstable network, which place the vertebral segments with high values of fracture loads (peaks) as well as the adjacent segments at risk of VCF. For instance, the high fracture load at T11 places T10, T11 and T12 at risk of fracture. Accordingly, T12 has already fractured, and T10 and T11 remain at risk. The relative changes between adjacent vertebrae segments that indicate instability (extremely high fracture load values) enables ease of identification of segments at high fracture risk. Clinicians would be able to work with pre-emptive treatment strategies in future as they can focus on more targeted therapy options at the high-risk vertebrae segments [3]

Introduction. Osteoarthritis (OA) is a slow progressive disease and a huge economic burden. A new target for therapy could be a growth factor treatment to prevent the loss of cartilage following injuries to the joint. BMP-7 is a promising candidate for such a novel therapy based on growth factors. In this study we combined the chondroprotective effects of BMP-7 with a novel thermosensitive hydrogel to prevent cartilage degeneration in a murine OA model. M&M. A BDI based thermosensitive hydrogel (Pluronic 123 with Butandiisyocyanate (BDI); LivImplant GmbH, Germany) was augmented with BMP-7 (rh-BMP-7, Olympus Biotech, France; 0.2 µg BMP-7/10µg Hydroge). To investigate the effects on OA progression we used the murine DMM (Destabilization of the medial meniscus) model for OA induction. Animal testing was approved by the Government Commitee of Upper Bavaria (file reference: 55.2-1-54-2532-150-13). A total of 38 C57BL/6 mice were included in this study. Immediately after the DMM surgery and wound closure BMP-7 mixed with BDI Hydrogel or only the BDI Hydrogel was administered via intraarticular injection. The following groups were examined: A) BMP-7 augmented BDI hydrogel B) only BDI hydrogel C) no injection following surgery D) control, healthy contralateral knee joint. After 4 (n=4 per group) and 8 (n=8) weeks mice were euthanized and knees were compared histologically. Results/Discussion. After 4 weeks the BMP-7 treated group showed a significant lower cartilage erosion compared to the group which only received DMM surgery. In the BMP-7 treated knee, osteoarthritis progression was also milder after 8 weeks than in knees of the DMM group. In all knees, except the control group, cartilage degeneration further progressed throughout the observation period. The contralateral joints showed no severe OA. We did not observe any inflammation or systemic reaction to the hydrogel. Taken together, we can conclude that BMP-7 showed a positive effect on the cartilage structure. Yet, the effect of a single administration is not strong enough to see a significant effect after 8 weeks. Furthermore, we can conclude, that the intraarticular administration of a thermosensitive hydrogel is an easy and feasible way to administer active agents precise to the joint

Successful treatment of periprosthetic joint infection involves surgical intervention and identification of infecting organisms to enable targeted antibiotic therapy. Current guidelines recommend intra-operative culture sampling to include at least 4 tissue samples and for each sample to be taken with a separate instrument. We aimed to review current revision arthroplasty practice for Greater Glasgow, specifically comparing intra-operative sampling technique for infected revision cases with these guidelines. We reviewed the clinical notes of all patients undergoing lower limb revision arthroplasty procedures in Greater Glasgow Hospitals (WIG, GRI, SGH) from July 2013 to August 2014. Demographics of all cases were collected. For revision procedures performed for infection we recorded details of intraoperative samples taken (number, type and sampling technique) and time for samples to reach the laboratory. Results of microbiology cultures were reviewed. Two hundred and fifty five revision arthroplasty procedures (152 hips, 103 knees) were performed in the 12 month study period. Of these 57 (22%) were infected cases (28 hips, 29 knees). These cases were treated by 14 arthroplasty surgeons with a median number of 3 infected cases managed per surgeon (range 1–11). 58% of cases had the recommended number of tissue samples taken. The median number of microbiology samples collected was 4 (range 1–14). Most procedures (91%) had no documentation of whether separate instruments were used for sampling. Number of tissue samples taken (≥4, p=0.01), time to lab (<24 hours, p=0.03) were significantly associated with positive culture results. In Greater Glasgow, a large number of surgeons manage infected arthroplasty cases with variability in intra-operative sampling techniques. Sample collection adheres to guideline recommendations in 58% cases. Adhering to guideline standards increases the likelihood of positive tissue cultures. Implementation of a standardised approach to intra-operative sampling for infected cases may improve patient management