Two-day-treatment of Activin-A leads to transient change in SV-HFO osteoblast gene expression and reduction in matrix mineralization

Activins regulate bone formation by controlling osteoclasts and osteoblasts. We investigated Activin‐A mechanism of action on human osteoblast mineralization, RNA and microRNA (miRNA) expression profile. A single 2‐day treatment of Activin‐A at Day 5 of osteoblast differentiation significantly reduced matrix mineralization. Activin A‐treated osteoblasts responded with transient change in gene expression, in a 2‐wave‐fashion. The 38 genes differentially regulated during the first wave (within 8 hr after Activin A start) were involved in transcription regulation. In the second wave (1–2 days after Activin A start), 65 genes were differentially regulated and related to extracellular matrix. Differentially expressed genes in both waves were associated to transforming growth factor beta signaling. We identified which microRNAs modulating osteoblast differentiation were regulated by Activin‐A. In summary, 2‐day treatment with Activin‐A in premineralization period of osteoblast cultures influenced miRNAs, gene transcription, and reduced matrix mineralization. Modulation of Activin A signaling might be useful to control bone quality for therapeutic purposes

Understanding Age-Induced Cortical Porosity in Women: The Accumulation and Coalescence of Eroded Cavities Upon Existing Intracortical Canals Is the Main Contributor

Intracortical bone remodeling normally ensures maintenance of the cortical bone matrix and strength, but during aging, this remodeling generates excessive porosity. The mechanism behind the age-induced cortical porosity is poorly understood and addressed in the present study. This study consists of a histomorphometric analysis of sections of iliac bone specimens from 35 women (age 16–78 years). First, the study shows that the age-induced cortical po

Recellularization of auricular cartilage via elastase-generated channels

Decellularized tissue matrices are promising substrates for tissue generation by stem cells to replace poorly regenerating tissues such as cartilage. However, the dense matrix of decellularized cartilage impedes colonisation by stem cells. Here, we show that digestion of elastin fibre bundles traversing auricular cartilage creates channels through which cells can migrate into the matrix. Human chondrocytes and bone marrow-derived mesenchymal stromal cells efficiently colonise elastin-treated scaffolds through these channels, restoring a glycosaminoglycan-rich matrix and improving mechanical properties while maintaining size and shape of the restored tiss

Hydroxychloroquine decreases human MSC-derived osteoblast differentiation and mineralization in vitro

We recently showed that patients with primary Sjögren Syndrome (pSS) have significantly higher bone mineral density (BMD) compared to healthy controls. The majority of those patients (69%) was using hydroxychloroquine (HCQ), which may have favourable effects on BMD. To study the direct effects of HCQ on human MSC-derived osteoblast activity. Osteoblasts were cultured from human mesenchymal stromal cells (hMSCs). Cultures were treated with different HCQ doses (control, 1 and 5 µg/ml). Alkaline phosphatase activity and calcium measurements were performed to evaluate osteoblast differentiation and activity, respectively. Detailed microarray analysis was performed in 5 µg/ml HCQ-treated cells and controls followed by qPCR validation. Additional cultures were performed using the cholesterol synthesis inhibitor simvastatin (SIM) to evaluate a potential mechanism of action. We showed that HCQ inhibits both MSC-derived osteoblast differentiation and mineralization in vitro. Microarray analysis and additional PCR validation reveale

1α,25-(OH)2D3 acts in the early phase of osteoblast differentiation to enhance mineralization via accelerated production of mature matrix vesicles

1α,25-dihydroxyitamin D3 (1,25D3) deficiency leads to impaired bone mineralization. We used the human pre-osteoblastic cell line SV-HFO, which forms within 19 days of culture an extracellular matrix that starts to mineralize around day 12, to examine th

Human mesenchymal stromal cells in adhesion to cell-derived extracellular matrix and titanium: Comparative kinome profile analysis

The extracellular matrix (ECM) physically supports cells and influences stem cell behaviour, modulating kinase-mediated signalling cascades. Cell-derived ECMs have emerged in bone regeneration as they reproduce physiological tissue-architecture and ameliorate mesenchymal stromal cell (MSC) properties. Titanium scaffolds show good mechanical properties, facilitate cell adhesion, and have been routinely used for bone tissue engineering (BTE). We analyzed the kinomic signature of human MSCs in adhesion to an osteopromotive osteoblast-derived ECM, and compared it to MSCs on titanium. PamChip kinase-array analysis revealed 63 phosphorylated peptides on ECM and 59 on titanium, with MSCs on ECM exhibiting significantly higher kinase activity than on titanium. MSCs on the two substrates showed overlapping kinome profiles, with activation of similar signalling pathways (FAK, ERK, and PI3K signalling). Inhibition of PI3K signalling in cells significantly reduced adhesion to ECM and increased the number of nonadherent cells on both substrates. In summary, this study comprehensively characterized the kinase activity in MSCs on cell-derived ECM and titanium, highlighting the role of PI3K signalling in kinomic changes regulating osteoblast viability and adhesion. Kinome profile analysis represents a powerful tool to select pathways to better understand cell behaviour. Osteoblast-derived ECM could be further investigated as titanium scaffold-coating to improve BTE

Skin Autofluorescence, a Noninvasive Biomarker for Advanced Glycation End-Products, Is Associated With Prevalent Vertebral and Major Osteoporotic Fractures: The Rotterdam Study

Advanced glycation end-products (AGEs), which bind to type 1 collagen in bone and skin, have been implicated in reduced bone quality. The AGE reader™ measures skin autofluorescence (SAF), which might be regarded as a marker of long-term accumulation of AGEs in tissues. We investigated the association of SAF with bone mineral density (BMD) and fractures in the general population. We studied 2853 individuals from the Rotterdam Study with available SAF measurements (median age, 74.1 years) and with data on prevalent major osteoporotic (MOFs: hip, humerus, wrist, clinical vertebral) and vertebral fractures (VFs: clinical + radiographic Genant’s grade 2 and 3). Radiographs were assessed 4 to 5 years before SAF. Multivariate regression models were performed adjusted for age, sex, BMI, creatinine, smoking status, and presence of diabetes and additionally for BMD with interaction terms to test for effect modification. Prevalence of MOFs was 8.5% and of VFs 7%. SAF had a curvilinear association with prevalent MOFs and VFs and therefore, age-adjusted, sex stratified SAF quartiles were used. The odds ratio (OR) (95% confidence interval [CI]) of the second, third and fourth quartiles of SAF for MOFs were as follows: OR 1.60 (95% CI, 1.08–2.35; p = .02); OR 1.30 (95% CI, 0.89–1.97; p = .20), and OR 1.40 (95% CI, 0.95–2.10; p = .09), respectively, with first (lowest) quartile as reference. For VFs the ORs were as follows: OR 1.69 (95% CI, 1.08–2.64; p = .02), OR 1.74(95% CI, 1.11–2.71; p = .01), and OR 1.73 (95% CI, 1.12–2.73; p = .02) for second, third, and fourth quartiles, respectively. When compari

Zika virus infection perturbs osteoblast function

Zika virus (ZIKV) infection is typically characterized by a mild self-limiting disease presenting with fever, rash, myalgia and arthralgia and severe fetal complications during pregnancy such as microcephaly, subcortical calcifications and arthrogyropsis. Virus-induced arthralgia due to perturbed osteoblast function has been described for other arboviruses. In case of ZIKV infection, the role of osteoblasts in ZIKV pathogenesis and bone related pathology remains unknown. Here, we study the effect of ZIKV infection on osteoblast differentiation, maturation and function by quantifying activity and gene expression of key biomarkers, using human bone marrow-derived mesenchymal stromal cells (MSCs, osteoblast precursors). MSCs were induced to differentiate into osteoblasts and we found that osteoblasts were highly susceptible to ZIKV infection. While infection did not cause a cytopathic effect, a significant reduction of key osteogenic markers such as ALP, RUNX2, calcium contents and increased expression of IL6 in ZIKV-infected MSCs implicated a delay in osteoblast development and maturation, as compared to uninfected controls. In conclusion, we have developed and characterized a new in vitro model to study the role of bone development in ZIKV pathogenesis, which will help to identify possible new targets for developing therapeutic and preventive measures

Osteoblasts secrete miRNA-containing extracellular vesicles that enhance expansion of human umbilical cord blood cells

Osteolineage cells represent one of the critical bone marrow niche components that support maintenance of hematopoietic stem and progenitor cells (HSPCs). Recent studies demonstrate that extracellular vesicles (EVs) regulate stem cell development via horizontal transfer of bioactive cargo, including microRNAs (miRNAs). Using next-generation sequencing we show that human osteoblast-derived EVs contain highly abundant miRNAs specifically enriched in EVs, including critical regulators of hematopoietic proliferation (e.g., miR-29a). EV treatment of human umbilical cord blood-derived CD34 + HSPCs alters the expression of candidate miRNA targets, such as HBP1, BCL2 and PTEN. Furthermore, EVs enhance proliferation of CD34 + cells and their immature subsets in growth factor-driven ex vivo expansion cultures. Importantly, EV-expanded cells retain their differentiation capacity in vitro and successfully engraft in vivo. These discoveries reveal a novel osteoblast-derived EV-mediated mechanism for regulation of HSPC proliferation and warrant consideration of EV-miRNAs for the development of expansion strategies to treat hematological disorders

Submicron patterns-on-a-chip: Fabrication of a microfluidic device incorporating 3D printed surface ornaments

Manufacturing high throughput in vitro models resembling the tissue microenvironment is highly demanded for studying bone regeneration. Tissues such as bone have complex multiscale architectures insid