Micro-computed tomography reconstructions of tibiae of stem cell transplanted osteogenesis imperfecta mice

Micro-computed tomography (micro-CT) is commonly used to assess bone quality and to evaluate the outcome of experimental therapies in animal models of bone diseases. Generating large datasets is however challenging and data are rarely made publicly available through shared repositories. Here we describe a dataset of micro-CT reconstructed scans of the proximal part of 21 tibiae from wild-type mice, osteogenesis imperfecta mice (homozygous oim/oim) and oim/oim mice transplanted with human amniotic fluid stem cells. The dataset contains, for each sample, 991 8-bit Bitmap reconstructed images and a 3D reconstruction of the bone in the PLY format, available at the online repository Figshare. In line with the increasing effort to make scientific datasets open-access, our data can be downloaded and used by other researchers to compare their observations with ours and to directly test scientific questions on osteogenesis imperfecta bones without the need to generate complete datasets

Modulation of osteoblast differentiation and function by the P2X4 receptor

Bone cells are known to express multiple P2 receptor subtypes, and the functional effects of receptor activation have been described for many of these. One exception is the P2X4 receptor, which despite strong expression in osteoblasts and osteoclasts, has no defined functional activity. This study used the selective P2X4 receptor antagonists, 5-BDBD and PSB-12062, to investigate the role of this receptor in bone. Both antagonists (≥ 0.1 μM) dose-dependently decreased bone formation by 60–100%. This was accompanied by a ≤ 70% decrease in alkaline phosphatase activity, a ≤ 40% reduction in cell number, and a ≤ 80% increase in the number of adipocytes present in the culture. The analysis of gene expression showed that levels of osteoblast marker genes (e.g. Alpl, Bglap) were decreased in 5-BDBD treated cells. Conversely, expression of the adipogenic transcription factor PPARG was increased 10-fold. In osteoclasts, high doses of both antagonists were associated with a reduction in osteoclast formation and resorptive activity by ≤ 95% and ≤ 90%, respectively. Taken together, these data suggest that the P2X4 receptor plays a role in modulating bone cell function. In particular, it appears to influence osteoblast differentiation favouring the osteogenic lineage over the adipogenic lineage

A machine learning‑based image segmentation method to quantify in vitro osteoclast culture endpoints

Quantification of in vitro osteoclast cultures (e.g. cell number) often relies on manual counting methods. These approaches are labour intensive, time consuming and result in substantial inter- and intra-user variability. This study aimed to develop and validate an automated workflow to robustly quantify in vitro osteoclast cultures. Using ilastik, a machine learning-based image analysis software, images of tartrate resistant acid phosphatase-stained mouse osteoclasts cultured on dentine discs were used to train the ilastik-based algorithm. Assessment of algorithm training showed that osteoclast numbers strongly correlated between manual- and automatically quantified values (r = 0.87). Osteoclasts were consistently faithfully segmented by the model when visually compared to the original reflective light images. The ability of this method to detect changes in osteoclast number in response to different treatments was validated using zoledronate, ticagrelor, and co-culture with MCF7 breast cancer cells. Manual and automated counting methods detected a 70% reduction (p < 0.05) in osteoclast number, when cultured with 10 nM zoledronate and a dose-dependent decrease with 1-10 μM ticagrelor (p < 0.05). Co-culture with MCF7 cells increased osteoclast number by ≥ 50% irrespective of quantification method. Overall, an automated image segmentation and analysis workflow, which consistently and sensitively identified in vitro osteoclasts, was developed. Advantages of this workflow are (1) significantly reduction in user variability of endpoint measurements (93%) and analysis time (80%); (2) detection of osteoclasts cultured on different substrates from different species; and (3) easy to use and freely available to use along with tutorial resources

Fabrication and characterization of Ag- and Ga-doped mesoporous glass-coated scaffolds based on natural marine sponges with improved mechanical properties

Natural marine sponges were used as sacrificial template for the fabrication of bioactive glassbased scaffolds. After sintering at 1050 ºC, the resulting samples were additionally coated with a sol silicate solution containing biologically active ions (Ag and Ga), well-known for their antibacterial properties in comparison with standard scaffolds made by PU foam templates. The produced scaffolds were characterized by superior mechanical properties (maximum compressive strength of 4 MPa) and total porosity of ~80%. Direct cell culture tests performed on the uncoated and coated samples showed positive results in terms of adhesion, proliferation, and differentiation of MC3T3-E1 cells. Moreover, vascular endothelial growth factor (VEGF) secretion from cells in contact with scaffold dissolution products was measured after 7 and 10 days of incubation, showing promising angiogenic results for bone tissue engineering applications. The antibacterial potential of the produced samples was assessed by performing agar diffusion tests against both Gram-positive and Gram-negative bacteria.EU Horizon 2020 project COACH 64255

Sheep condyle model evaluation of bone marrow cell concentrate combined with a scaffold for repair of large osteochondral defects

AIMS: Minimally manipulated cells, such as autologous bone marrow concentrates (BMC), have been investigated in orthopaedics as both a primary therapeutic and augmentation to existing restoration procedures. However, the efficacy of BMC in combination with tissue engineering is still unclear. In this study, we aimed to determine whether the addition of BMC to an osteochondral scaffold is safe and can improve the repair of large osteochondral defects when compared to the scaffold alone. METHODS: The ovine femoral condyle model was used. Bone marrow was aspirated, concentrated, and used intraoperatively with a collagen/hydroxyapatite scaffold to fill the osteochondral defects (n = 6). Tissue regeneration was then assessed versus the scaffold-only group (n = 6). Histological staining of cartilage with alcian blue and safranin-O, changes in chondrogenic gene expression, microCT, peripheral quantitative CT (pQCT), and force-plate gait analyses were performed. Lymph nodes and blood were analyzed for safety. RESULTS: The results six months postoperatively showed that there were no significant differences in bone regrowth and mineral density between BMC-treated animals and controls. A significant upregulation of messenger RNA (mRNA) for types I and II collagens in the BMC group was observed, but there were no differences in the formation of hyaline-like cartilage between the groups. A trend towards reduced sulphated glycosaminoglycans (sGAG) breakdown was detected in the BMC group but this was not statistically significant. Functional weightbearing was not affected by the inclusion of BMC. CONCLUSION: Our results indicated that the addition of BMC to scaffold is safe and has some potentially beneficial effects on osteochondral-tissue regeneration, but not on the functional endpoint of orthopaedic interest. Cite this article: Bone Joint Res 2021;10(10):677-689

Bioengineering the ameloblastoma tumour to study its effect on bone nodule formation

Ameloblastoma is a benign, epithelial cancer of the jawbone, which causes bone resorption and disfigurement to patients affected. The interaction of ameloblastoma with its tumour stroma drives invasion and progression. We used stiff collagen matrices to engineer active bone forming stroma, to probe the interaction of ameloblastoma with its native tumour bone microenvironment. This bone-stroma was assessed by nano-CT, transmission electron microscopy (TEM), Raman spectroscopy and gene analysis. Furthermore, we investigated gene correlation between bone forming 3D bone stroma and ameloblastoma introduced 3D bone stroma. Ameloblastoma cells increased expression of MMP-2 and -9 and RANK temporally in 3D compared to 2D. Our 3D biomimetic model formed bone nodules of an average surface area of 0.1 mm2 and average height of 92.37 ± 7.96 μm over 21 days. We demonstrate a woven bone phenotype with distinct mineral and matrix components and increased expression of bone formation genes in our engineered bone. Introducing ameloblastoma to the bone stroma, completely inhibited bone formation, in a spatially specific manner. Multivariate gene analysis showed that ameloblastoma cells downregulate bone formation genes such as RUNX2. Through the development of a comprehensive bone stroma, we show that an ameloblastoma tumour mass prevents osteoblasts from forming new bone nodules and severely restricted the growth of existing bone nodules. We have identified potential pathways for this inhibition. More critically, we present novel findings on the interaction of stromal osteoblasts with ameloblastoma

Rheumatoid synovial fluid interleukin-17-producing CD4 T cells have abundant tumor necrosis factor-alpha co-expression, but little interleukin-22 and interleukin-23R expression

Introduction\ud Th17 cells have been implicated in the pathogenesis of rheumatoid arthritis (RA). The aim of this study was to systematically analyse the phenotype, cytokine profile and frequency of interleukin-17 (IL-17) producing CD4-positive T cells in mononuclear cells isolated from peripheral blood, synovial fluid and synovial tissue of RA patients with established disease, and to correlate cell frequencies with disease activity. \ud \ud Methods\ud Flow cytometry was used to analyse the phenotype and cytokine production of mononuclear cells isolated from peripheral blood (PBMC) (n = 44), synovial fluid (SFMC) (n = 14) and synovium (SVMC) (n = 10) of RA patients and PBMC of healthy controls (n = 13). \ud \ud Results\ud The frequency of IL-17-producing CD4 T cells was elevated in RA SFMC compared with RA PBMC (P = 0.04). However, the frequency of this population in RA SVMC was comparable to that in paired RA PBMC. The percentage of IL-17-producing CD4 T cells coexpressing tumor necrosis factor alpha (TNFα) was significantly increased in SFMC (P = 0.0068). The frequency of IFNγ-producing CD4 T cells was also significantly higher in SFMC than paired PBMC (P = 0.042). The majority of IL-17-producing CD4 T cells coexpressed IFNγ. IL-17-producing CD4 T cells in RA PBMC and SFMC exhibited very little IL-22 or IL-23R coexpression. \ud \ud Conclusions\ud These findings demonstrate a modest enrichment of IL-17-producing CD4 T cells in RA SFMC compared to PBMC. Th17 cells in SFMC produce more TNFα than their PBMC counterparts, but are not a significant source of IL-22 and do not express IL-23R. However, the percentage of CD4 T cells which produce IL-17 in the rheumatoid joint is low, suggesting that other cells may be alternative sources of IL-17 within the joints of RA patients. \ud \u

Effect of a supplement rich in alkaline minerals on acid-base balance in humans

This is an Open Access article distributed under the terms of the Creative Commons Attribution Licens

Consumer perceptions of co-branding alliances: Organizational dissimilarity signals and brand fit

This study explores how consumers evaluate co-branding alliances between dissimilar partner firms. Customers are well aware that different firms are behind a co-branded product and observe the partner firms’ characteristics. Drawing on signaling theory, we assert that consumers use organizational characteristics as signals in their assessment of brand fit and for their purchasing decisions. Some organizational signals are beyond the control of the co-branding partners or at least they cannot alter them on short notice. We use a quasi-experimental design and test how co-branding partner dissimilarity affects brand fit perception. The results show that co-branding partner dissimilarity in terms of firm size, industry scope, and country-of-origin image negatively affects brand fit perception. Firm age dissimilarity does not exert significant influence. Because brand fit generally fosters a benevolent consumer attitude towards a co-branding alliance, the findings suggest that high partner dissimilarity may reduce overall co-branding alliance performance

Association between AIRE gene polymorphism and rheumatoid arthritis: a systematic review and meta-analysis of case-control studies.

Autoimmune regulator (AIRE) is a transcription factor that functions as a novel player in immunological investigations. In the thymus, it has a pivotal role in the negative selection of naive T-cells during central tolerance. Experimental studies have shown that single nucleotide polymorphism (SNP) alters transcription of the AIRE gene. SNPs thereby provide a less efficient negative selection, propagate higher survival of autoimmune T-cells, and elevate susceptibility to autoimmune diseases. To date, only rheumatoid arthritis (RA) has been analysed by epidemiological investigations in relation to SNPs in AIRE. In our meta-analysis, we sought to encompass case-control studies and confirm that the association between SNP occurrence and RA. After robust searches of Embase, PubMed, Cochrane Library, and Web of Science databases, we found 19 articles that included five independent studies. Out of 11 polymorphisms, two (rs2075876, rs760426) were common in the five case-control studies. Thus, we performed a meta-analysis for rs2075876 (7145 cases and 8579 controls) and rs760426 (6696 cases and 8164 controls). Our results prove that rs2075876 and rs760426 are significantly associated with an increased risk of RA in allelic, dominant, recessive, codominant heterozygous, and codominant homozygous genetic models. These findings are primarily based on data from Asian populations