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Introduction Rheumatoid arthritis (RA) is the most common inflammatory, erosive polyarthritis and affects approximately 0.5%-1.5% of the worldwide population. RA is characterized by breach of self-tolerance and production of several [...], Ectopic lymphoid structures (ELSs) in the rheumatoid synovial joints sustain autoreactivity against locally expressed autoantigens. We recently identified recombinant monoclonal antibodies (RA- rmAbs) derived from single, locally differentiated rheumatoid arthritis (RA) synovial B cells, which specifically recognize fibroblast-like synoviocytes (FLSs). Here, we aimed to identify the specificity of FLS-derived autoantigens fueling local autoimmunity and the functional role of anti-FLS antibodies in promoting chronic inflammation. A subset of anti-FLS RA- rmAbs reacting with a 60 kDa band from FLS extracts demonstrated specificity for HSP60 and partial cross-reactivity to other stromal autoantigens (i.e., calreticulin/vimentin) but not to citrullinated fibrinogen. Anti-FLS RA-rmAbs, but not anti-neutrophil extracellular traps rmAbs, exhibited pathogenic properties in a mouse model of collagen-induced arthritis. In patients, anti-HSP60 antibodies were preferentially detected in RA versus osteoarthritis (OA) synovial fluid. Synovial HSPD1 and CALR gene expression analyzed using bulk RNA-Seq and GeoMx-DSP closely correlated with the lympho-myeloid RA pathotype, and HSP60 protein expression was predominantly observed around ELS. Moreover, we observed a significant reduction in synovial HSP60 gene expression followed B cell depletion with rituximab that was strongly associated with the treatment response. Overall, we report that synovial stromal-derived autoantigens are targeted by pathogenic autoantibodies and are associated with specific RA pathotypes, with potential value for patient stratification and as predictors of the response to B cell-depleting therapies.

Crouzon syndrome is a congenital craniofacial disorder caused by mutations in the Fibroblast Growth Factor Receptor 2 (FGFR2). It is characterized by the premature fusion of cranial sutures, leading to a brachycephalic head shape, and midfacial hypoplasia. The aim of this study was to investigate the effect of the FGFR2 mutation on the microarchitecture of cranial bones at different stages of postnatal skull development, using the FGFR2C342Y mouse model. Apart from craniosynostosis, this model shows cranial bone abnormalities. High‐resolution synchrotron microtomography images of the frontal and parietal bone were acquired for both FGFR2C342Y/+ (Crouzon, heterozygous mutant) and FGFR2+/+ (control, wild‐type) mice at five ages (postnatal days 1, 3, 7, 14 and 21, n = 6 each). Morphometric measurements were determined for cortical bone porosity: osteocyte lacunae and canals. General linear model to assess the effect of age, anatomical location and genotype was carried out for each morphometric measurement. Histological analysis was performed to validate the findings. In both groups (Crouzon and wild‐type), statistical difference in bone volume fraction, average canal volume, lacunar number density, lacunar volume density and canal volume density was found at most age points, with the frontal bone generally showing higher porosity and fewer lacunae. Frontal bone showed differences between the Crouzon and wild‐type groups in terms of lacunar morphometry (average lacunar volume, lacunar number density and lacunar volume density) with larger, less dense lacunae around the postnatal age of P7–P14. Histological analysis of bone showed marked differences in frontal bone only. These findings provide a better understanding of the pathogenesis of Crouzon syndrome and will contribute to computational models that predict postoperative changes with the aim to improve surgical outcome. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is characterised by the deterioration of cartilage in the joints and pain. We hypothesise that semaphorin‐3A (sema‐3A), a chemorepellent for sensory nerves, plays a role in joint degradation and pain. We used the mechanical joint loading (MJL) model of OA to investigate sema‐3A expression in the joint and examine its association with the development of OA and pain. We also analyse its effect on chondrocyte differentiation using the ATDC5 cell line. We demonstrate that sema‐3A is present in most tissues in the healthy joint and its expression increases in highly innervated tissues, such as cruciate ligaments, synovial lining and subchondral bone, in loaded compared to nonloaded control joints. In contrast, sema‐3A expression in cartilage was decreased in the severe OA induced by the application of high loads. There was a significant increase in circulating sema‐3A, 6 weeks after MJL compared to the nonloaded mice. mRNA for sema‐3A and its receptor Plexin A1 were upregulated in the dorsal root ganglia of mice submitted to MJL. These increases were supressed by zoledronate, an inhibitor of bone pain. Sema‐3A was expressed at all stages of Chondrocyte maturation and, when added exogenously, stimulated expression of markers of chondrocyte differentiation. This indicates that sema‐3A could affect joint tissues distinctively during the development of OA. In highly innervated joint tissues, sema‐3A could control innervation and/or induce pain‐associated neuronal changes. In cartilage, sema‐3A could favour its degeneration by modifying chondrocyte differentiation. Significance statement: Semaphorin‐3A (sema‐3A) is an axon guidance molecule previously shown to play a role in neural ingrowth and vascularisation during degeneration of tissues. We investigated its expression in tissues of the mouse joint and examined changes in expression with the development of osteoarthritis (OA). We show that sema‐3A expression is increased in highly innervated joint tissues with OA and may control innervation in these tissues. In contrast, sema‐3A expression in cartilage decreases with the severity of OA. We also demonstrate using a chondrocytic cell line that sema‐3A stimulates expression of markers of chondrocyte differentiation and may play a role in cartilage degeneration. [ABSTRACT FROM AUTHOR]

The administration of intermittent parathyroid hormone (iPTH) is anabolic to the skeleton. Recent studies with cultured osteoblasts have revealed that the expression of PHOSPHO1, a bone‐specific phosphatase essential for the initiation of mineralisation, is regulated by PTH. Therefore, this study sought to determine whether the bone anabolic response to iPTH involves modulation of expression of Phospho1 and of other enzymes critical for bone matrix mineralisation. To mimic iPTH treatment, primary murine osteoblasts were challenged with 50 nM PTH for 6 h in every 48 h period for 8 days (4 cycles), 14 days (7 cycles) and 20 days (10 cycles) in total. The expression of both Phospho1 and Smpd3 was almost completely inhibited after 4 cycles, whereas 10 cycles were required to stimulate a similar response in Alpl expression. To explore the in vivo role of PHOSPHO1 in PTH‐mediated osteogenesis, the effects of 14‐ and 28‐day iPTH (80 µg/kg/day) administration was assessed in male wild‐type (WT) and Phospho1−/− mice. The expression of Phospho1, Alpl, Smpd3, Enpp1, Runx2 and Trps1 expression was enhanced in the femora of WT mice following iPTH administration but remained unchanged in the femora of Phospho1−/− mice. After 28 days of iPTH administration, the anabolic response in the femora of WT was greater than that noted in Phospho1−/− mice. Specifically, cortical and trabecular bone volume/total volume, as well as cortical thickness, were increased in femora of iPTH‐treated WT but not in iPTH‐treated Phospho1−/− mice. Trabecular bone osteoblast number was also increased in iPTH‐treated WT mice but not in iPTH‐treated Phospho1−/− mice. The increased levels of Phospho1, Alpl, Enpp1 and Smpd3 in WT mice in response to iPTH administration is consistent with their contribution to the potent anabolic properties of iPTH in bone. Furthermore, as the anabolic response to iPTH was attenuated in mice deficient in PHOSPHO1, this suggests that the osteoanabolic effects of iPTH are at least partly mediated via bone mineralisation processes. Significance statement: Intermittent administration of parathyroid hormone (iPTH) is recognised to increase osteoblast number and promote bone formation. A critical component of this anabolic response is the mineralisation of the osteoid matrix but the effects iPTH has on the expression of mineralisation regulating enzymes is unclear. This study's principal finding was that iPTH administration increased the expression of Phospho1, Alpl, Enpp1 and Smpd3 in vivo which was consistent with a bone anabolic PTH regimen. The data also disclosed that in the absence of PHOSPHO1, the iPTH anabolic response was dampened, suggesting that amplified Phospho1 expression is a prerequisite for a full iPTH‐mediated bone anabolic response. [ABSTRACT FROM AUTHOR]

Despite knowledge that sexually dimorphic mechanisms regulate bone homeostasis, sex often remains unreported and unconsidered in preclinical experimental design. Failure to report sex could lead to inappropriate generalizations of research findings and less effective translation into clinical practice. Preclinical sex bias (preferential selection of one sex) is present across other fields, including neuroscience and immunology, but remains uninvestigated in skeletal research. For context, we first summarized key literature describing sexually dimorphic bone phenotypes in mice. We then investigated sex reporting practices in skeletal research, specifically how customary it is for murine sex to be included in journal article titles or abstracts and then determined whether any bias in sex reporting exists. Because sex hormones are important regulators of bone health (gonadectomy procedures, ie, ovariectomy [OVX] and orchidectomy [ORX], are common yet typically not reported with sex), we incorporated reporting of OVX and ORX terms, representing female and male mice, respectively, into our investigations around sex bias. Between 1999 and 2020, inclusion of sex in titles or abstracts was low in murine skeletal studies (2.6%–4.06%). Reporting of OVX and ORX terms was low (1.44%–2.64%) and reporting of OVX and ORX with sex uncommon (0.4%–0.3%). When studies were combined to include both sexes and OVX (representing female) and ORX terms (representing male), a bias toward reporting of female mice was evident. However, when the terms OVX and ORX were removed, a bias toward the use of male mice was identified. Thus, studies focusing on sex hormones are biased toward female reporting with all other studies biased in reporting of male mice. We now call upon journal editors to introduce consistent guidance for transparent and accessible reporting of murine sex in skeletal research to better monitor preclinical sex bias, to diversify development of treatments for bone health, and to enable global skeletal health equity. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR). [ABSTRACT FROM AUTHOR]

Multiple prevalent diseases, such as osteoarthritis (OA), for which there is no cure or full understanding, affect the osteochondral unit; a complex interface tissue whose architecture, mechanical nature and physiological characteristics are still yet to be successfully reproduced in vitro. Although there have been multiple tissue engineering-based approaches to recapitulate the three dimensional (3D) structural complexity of the osteochondral unit, there are various aspects that still need to be improved. This review presents the different pre-requisites necessary to develop a human osteochondral unit construct and focuses on 3D bioprinting as a promising manufacturing technique. Examples of 3D bioprinted osteochondral tissues are reviewed, focusing on the most used bioinks, chosen cell types and growth factors. Further information regarding the applications of these 3D bioprinted tissues in the fields of disease modelling, drug testing and implantation is presented. Finally, special attention is given to the limitations that currently hold back these 3D bioprinted tissues from being used as models to investigate diseases such as OA. Information regarding improvements needed in bioink development, bioreactor use, vascularisation and inclusion of additional tissues to further complete an OA disease model, are presented. Overall, this review gives an overview of the evolution in 3D bioprinting of the osteochondral unit and its applications, as well as further illustrating limitations and improvements that could be performed explicitly for disease modelling. [ABSTRACT FROM AUTHOR]

Patients with advanced chronic kidney disease (CKD) often present with skeletal abnormalities, a condition known as renal osteodystrophy (ROD). While tissue nonspecific alkaline phosphatase (TNAP) and PHOSPHO1 are critical f or bone mineralization, their role in the etiology of ROD is unclear. To address this, ROD was induced in both WT and Phospho1 knockout (P1KO) mice through dietary adenine supplementation. The mice presented with hyperphosphatemia, hyperparathyroidism, and elevated levels of FGF23 and bone turnover markers. In particular, we noted that in CKD mice, bone mineral density (BMD) was increased in cortical bone (P < 0.05) but decreased in trabecular bone (P < 0.05). These changes were accompanied by decreased TNAP ( P < 0.01) and increased PHOSPHO1 (P < 0.001) expression in WT CKD bones. In P1KO CKD mice, the cortical BMD phenotype was rescued, suggesting that the increased cortical BMD of CKD mice was driven by increased PHOSPHO1 expression. Ot her structural parameters were also improved in P1KO CKD mice. We further investigated the driver of the mineralization defects, by studying the effects of FGF23, PTH, and phosphate administration on PHOSPHO1 and TNAP expression by primary murin e osteoblasts. We found both PHOSPHO1 and TNAP expressions to be downregulated in response to phosphate and PTH. The in vitro data suggest that the TNAP reduction in CKD-MBD is driven by the hyperphosphatemia and/or hyperparathyroidism noted in these mice, while the higher PHOSPHO1 expression may be a compensatory mech anism. Increased PHOSPHO1 expression in ROD may contribute to the disordered ske letal mineralization characteristic of this progressive disorder. [ABSTRACT FROM AUTHOR]

Femoral neck fractures are a massive personal and health programme burden. Methods to study femoral neck strength, across its combined trabecular and cortical components are therefore essential. Rodent ovariectomy-induced osteoporosis models are commonly coupled with ex vivo 3-/4-point bending methods to measure changes in femoral cortical diaphysis. The loading direction used to assess these properties are often non-physiologic and, moreover, these ovariectomy models are linked to marked weight gain that can influence the biomechanical properties. Herein, we explore whether more physiological axial ex vivo loading protocols applied to femoral neck samples of ovariectomised (OVX) rodents provide anatomically-relevant models for the assessment of strength. We examine the use of mouse and rat femurs, loaded in constrained and unconstrained configuration, respectively, and explore whether weight-correction increases their utility. Accordingly, the mid-shaft of the proximal half of femurs from OVX and sham-operated (Sham) mice was methacrylate-anchored and the head loaded parallel to the diaphysis (constrained). Alternatively, femurs from OVX and Sham rats were isolated intact and axially-loaded through hip and knee joint articular surfaces (unconstrained). Yield displacement, stiffness, maximum load and resilience were measured and fracture pattern classified; effects of body weight-correction via a linear regression method or simple division were assessed. Our data reveal significant deficiencies in biomechanical properties in OVX mouse femurs loaded in constrained configuration, only after weight-correction by linear regression. In addition, evaluation of rat femur biomechanics in unconstrained loading demonstrated greater variation and that weight-correction by simple division improved scope to reveal significant OVX impact. We conclude that greater femoral neck fracture susceptibility can indeed be measured in OVX rodents as long as multiple biomechanical parameters are reported, care is taken in choosing the method for assessing load-bearing strength and weight-correction applied. These studies advance the establishment of more relevant rodent models for the study of femoral neck fracture. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a common chronic disease largely driven by mechanical factors, causing significant health and economic burdens worldwide. Early detection is challenging, making animal models a key tool for studying its onset and mechanically-relevant pathogenesis. This review evaluate current use of preclinical in vivo models and progressive measurement techniques for analysing biomechanical factors in the specific context of the clinical OA phenotypes. It categorizes preclinical in vivo models into naturally occurring, genetically modified, chemically-induced, surgically-induced, and non-invasive types, linking each to clinical phenotypes like chronic pain, inflammation, and mechanical overload. Specifically, we discriminate between mechanical and biological factors, give a new explanation of the mechanical overload OA phenotype and propose that it should be further subcategorized into two subtypes, post-traumatic and chronic overloading OA. This review then summarises the representative models and tools in biomechanical studies of OA. We highlight and identify how to develop a mechanical model without inflammatory sequelae and how to induce OA without significant experimental trauma and so enable the detection of changes indicative of early-stage OA in the absence of such sequelae. We propose that the most popular post-traumatic OA biomechanical models are not representative of all types of mechanical overloading OA and, in particular, identify a deficiency of current rodent models to represent the chronic overloading OA phenotype without requiring intraarticular surgery. We therefore pinpoint well standardized and reproducible chronic overloading models that are being developed to enable the study of early OA changes in non-trauma related, slowly-progressive OA. In particular, non-invasive models (repetitive small compression loading model and exercise model) and an extra-articular surgical model (osteotomy) are attractive ways to present the chronic natural course of primary OA. Use of these models and quantitative mechanical behaviour tools such as gait analysis and non-invasive imaging techniques show great promise in understanding the mechanical aspects of the onset and progression of OA in the context of chronic knee joint overloading. Further development of these models and the advanced characterisation tools will enable better replication of the human chronic overloading OA phenotype and thus facilitate mechanically-driven clinical questions to be answered., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Xu, Kazezian, Pitsillides and Bull.)