scholarly journals Extracellular matrix mineralization in the mouse osteoblast-like cell line MC3T3-E1 is regulated by actin cytoskeleton reorganization and non-protein molecules secreted from the cells themselves

2019 ◽  
Author(s):  
Hiraku Suzuki ◽  
Kazuaki Tatei ◽  
Noriyasu Ohshima ◽  
Seiichi Sato ◽  
Takashi Izumi
Keyword(s):  
Bone Formation ◽  
Actin Cytoskeleton ◽  
Mrna Expression ◽  
Cell Line ◽  
Late Stage ◽  
Matrix Mineralization ◽  
Middle Stage ◽  
Cytoskeleton Reorganization ◽  
Protein Molecules ◽  
Mouse Osteoblast

AbstractBone tissue constantly undergoes turnover via bone formation by osteoblasts and bone resorption by osteoclasts. This process enables bone to maintain its overall shape while altering its local structure. However, the detailed mechanism of how osteoblast cell-signaling systems induce various structural changes in bone tissue have not yet been completely elucidated. In this study, we focused on the actin cytoskeleton as a regulatory system for bone formation and constructed an in vitro experimental system using the mouse osteoblast-like cell line MC3T3-E1. We found that, in MC3T3-E1 cells, the actin cytoskeleton had an important role in matrix mineralization via activation of specific developmental pathways and it was regulated by non-protein molecules secreted from MC3T3-E1 cells themselves. In MC3T3-E1 cells, we observed changes of actin cytoskeleton reorganization and accumulation of PIP2 related to actin filament convergences during cell differentiation, in the undifferentiated, early, middle and late stage. Actin cytoskeleton disruption with Cyto D, polymerization inhibitor of actin filament, in early and middle stage cells induced significant increase of osteocalcin mRNA expression normally expressed only in late stage, decrease of Alkaline phosphatase mRNA expression after 24h and abnormal matrix mineralization in MC3T3-E1 cells. Inhibition of Giα with PTX known to regulate actin cytoskeleton in middle stage induced changes in the actin cytoskeleton and PIP2 accumulation and suppression of matrix mineralization after 5 days. Furthermore, addition of non-protein molecules from culture medium of cells at various differentiation stage induced difference of PIP2 accumulation after 5 min, actin cytoskeleton in 20 min, and matrix mineralization after 5 days. These results not only provide new knowledge about the actin cytoskeleton function in bone-forming cells, but also suggest that cell signaling via non-protein molecules such as lipids plays important roles in bone formation.

scholarly journals The influence of alliin on actin cytoskeleton reorganization in H1299 cell line in the presence of low doses of doxorubicin

2015 ◽  
Vol 29 (1) ◽  
pp. 25
Author(s):  
Agnieszka Glińska ◽  
Magdalena Izdebska ◽  
Marta Hałas ◽  
Anna Klimaszewska-Wiśniewska ◽  
Alina Grzanka
Keyword(s):  
Actin Cytoskeleton ◽  
Cell Line ◽  
Low Doses ◽  
H1299 Cell ◽  
Cytoskeleton Reorganization ◽  
H1299 Cell Line

scholarly journals Matrix Vesicles: Role in Bone Mineralization and Potential Use as Therapeutics

2021 ◽  
Vol 14 (4) ◽  
pp. 289
Author(s):  
Sana Ansari ◽  
Bregje W. M. de de Wildt ◽  
Michelle A. M. Vis ◽  
Carolina E. de de Korte ◽  
Keita Ito ◽  
...  
Keyword(s):  
Bone Formation ◽  
Bone Regeneration ◽  
Bone Cells ◽  
Bone Mineralization ◽  
Recipient Cell ◽  
Organic Matrix ◽  
Matrix Vesicles ◽  
Matrix Mineralization

Bone is a complex organ maintained by three main cell types: osteoblasts, osteoclasts, and osteocytes. During bone formation, osteoblasts deposit a mineralized organic matrix. Evidence shows that bone cells release extracellular vesicles (EVs): nano-sized bilayer vesicles, which are involved in intercellular communication by delivering their cargoes through protein–ligand interactions or fusion to the plasma membrane of the recipient cell. Osteoblasts shed a subset of EVs known as matrix vesicles (MtVs), which contain phosphatases, calcium, and inorganic phosphate. These vesicles are believed to have a major role in matrix mineralization, and they feature bone-targeting and osteo-inductive properties. Understanding their contribution in bone formation and mineralization could help to target bone pathologies or bone regeneration using novel approaches such as stimulating MtV secretion in vivo, or the administration of in vitro or biomimetically produced MtVs. This review attempts to discuss the role of MtVs in biomineralization and their potential application for bone pathologies and bone regeneration.

In Vivo and In Vitro Evaluation of Coated HAp Films with Different Surface Morphology

2007 ◽  
Vol 539-543 ◽  
pp. 710-715
Author(s):  
Kotaro Kuroda ◽  
Ryoichi Ichino ◽  
Masazumi Okido
Keyword(s):  
Bone Formation ◽  
Surface Morphology ◽  
New Bone Formation ◽  
Contact Ratio ◽  
Bone Implant ◽  
Ft Ir ◽  
Mouse Osteoblast ◽  
Surface Morphologies

Hydroxyapatite (HAp) coatings were formed on cp titanium plates and rods by the thermal substrate method in an aqueous solution that included 0.3 mM Ca(H2PO4)2 and 0.7 mM CaCl2. The coating experiments were conducted at 40-140 oC and pH = 8 for 15 or 30 min. The properties for the coated samples were studied using XRD, EDX, FT-IR, and SEM. All the specimens were covered with HAp, which had different surface morphologies such as net-like, plate-like and needle-like. After cleaning and sterilization, all the coated specimens were subjected to in vivo and vitro testing. In the in vitro testing, the mouse osteoblast-like cells (MC3T3-E1) were cultured on the coated and non-coated specimens for up to 30 days. Moreover, the specimens (φ2 x 5 mm) were implanted in rats femoral for up to 8 weeks, the osseoinductivity on them were evaluated. In in vitro evaluations, there were not significant differences between the different surface morphologies. In in vivo evaluations, however, two weeks postimplantation, new bone formed on both the HAp coated and non-coated titanium rods in the cancellous and cortical bone. The bone-implant contact ratio, which was used for the evaluation of new bone formation, was significantly dependent on the surface morphology of the HAp, and the results demonstrated that the needle-like coating appears to promote rapid bone formation.

CYTOKINE mRNA EXPRESSION IN HUMAN PLATELETS AND A MEGAKARYOCYTIC CELL LINE AND CYTOKINE MODULATION OF PLATELET FUNCTION

Cytokine ◽  
1997 ◽  
Vol 9 (6) ◽  
pp. 405-411 ◽  
Author(s):  
Gerald Soslau ◽  
Doris A. Morgan ◽  
Jonathan S. Jaffe ◽  
Isadore Brodsky ◽  
Yihe Wang
Keyword(s):  
Mrna Expression ◽  
Cell Line ◽  
Platelet Function ◽  
Human Platelets ◽  
Cytokine Mrna ◽  
Cytokine Modulation ◽  
Cytokine Mrna Expression ◽  
Megakaryocytic Cell

scholarly journals Induction of Colony-Stimulating Factor Expression following Staphylococcus or SalmonellaInteraction with Mouse or Human Osteoblasts

2000 ◽  
Vol 68 (9) ◽  
pp. 5075-5083 ◽  
Author(s):  
Kenneth L. Bost ◽  
Jennifer L. Bento ◽  
John K. Ellington ◽  
Ian Marriott ◽  
Michael C. Hudson
Keyword(s):  
Immune Response ◽  
Mrna Expression ◽  
Bone Diseases ◽  
Cytokine Secretion ◽  
Human Osteoblasts ◽  
Salmonella Dublin ◽  
Gm Csf ◽  
Common Causes ◽  
Mouse Osteoblast ◽  
Stimulating Factor

ABSTRACT Staphylococcus aureus and Salmonella spp. are common causes of bone diseases; however, the immune response during such infections is not well understood. Colony-stimulating factors (CSF) have a profound influence on osteoclastogenesis, as well as the development of immune responses following infection. Therefore, we questioned whether interaction of osteoblasts with two very different bacterial pathogens could affect CSF expression by these cells. Cultured mouse and human osteoblasts were exposed to various numbers ofS. aureus or Salmonella dublin bacteria, and a comprehensive analysis of granulocyte-macrophage (GM)-CSF, granulocyte (G)-CSF, macrophage (M)-CSF, and interleukin-3 (IL-3) mRNA expression and cytokine secretion was performed. Expression of M-CSF and IL-3 mRNAs by mouse osteoblasts was constitutive and did not increase significantly following bacterial exposure. In contrast, GM-CSF and G-CSF mRNA expression by mouse osteoblasts was dramatically upregulated following interaction with either viable S. aureus orSalmonella. This increased mRNA expression also translated into high levels of GM-CSF and G-CSF secretion by mouse and human osteoblasts following bacterial exposure. Viable S. aureusand Salmonella induced maximal levels of CSF mRNA expression and cytokine secretion compared to UV-killed bacteria. Furthermore, GM-CSF and G-CSF mRNA expression could be induced in unexposed osteoblasts separated by a permeable Transwell membrane from bacterially exposed osteoblasts. M-CSF secretion was increased in cultures of exposed human osteoblasts but not in exposed mouse osteoblast cultures. Together, these studies are the first to define CSF expression and suggest that, following bacterial exposure, osteoblasts may influence osteoclastogenesis, as well as the development of an immune response, via the production of these cytokines.

Abstract 017: Hyperglycemia-induced Posttranscriptional Modification Of Proinflammatory Cytokine Mrna Stability Is Mediated Via Hur/PKC-delta Signaling Pathway

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Suresh K Verma ◽  
Alexander R Mackie ◽  
Erin E Vaughan ◽  
Tatiana V Abramova ◽  
Raj kishore ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Cell Line ◽  
Signaling Pathway ◽  
Mrna Stability ◽  
Cardiac Fibrosis ◽  
Diabetic Patients ◽  
Tgf Beta ◽  
Intramyocardial Injection ◽  
Pkc Delta ◽  
Increased Risk

Patients with diabetes are predisposed to increased risk of cardiovascular diseases. Persistent interaction of infiltrating macrophages and resident fibroblasts play a critical role in cardiac fibrosis. However, the signaling mechanism is not clear. We hypothesized that macrophage ELAV1 (mRNA stabilizing protein) modulates profibrotic mediators and extracellular matrix turnover by binding to 3′UTR and regulating the mRNA stability of TGF-beta and MMP-9 in hyperglycemic conditions. Mice receiving intramyocardial injection of HuR-specific shRNA showed significant reduction in infarct size and fibrosis area. Reduced fibrosis was associated with decrease in TGF-beta and MMP-9 expression in the myocardium. Conditioned media (CM) from high glucose (HG) treated macrophages significantly increased profibrogenic response (increased mRNA expression of Col1a1, Col3a1 and fibronectin) in fibroblast cell line as compared to fibroblasts incubated with CM from low glucose (LG)-treated macrophages. Knockdown of ELAV1 in HG-treated macrophages abrogated the profibrotic effects in fibroblasts. Indirect immunofluroscence of bone marrow-derived macrophages (BMM) demonstrated that HG increases nuclear ELAV1 export to the cytoplasm. Pharmacological inhibition of Protein kinase C-delta (PKCd) blocked HG-induced ELAV1 nuclear to cytoplasmic translocation. In vitro, stable knockdown of ELAV1 in mouse macrophage cell line RAW 264.7 reduced mRNA expression of TGF-beta and MMP-9 following LPS challenge, accompanied by a marked reduction in the mRNA stability of these genes. Our study here establishes an ELAV1/TGF-beta/MMP-9/PKC-delta signaling axis in the macrophages controlling the profibrogenic responses in fibroblasts, the major contributor in the pathogenesis of fibrosis. Therefore, targeting this signaling pathway might be of therapy value for cardiac fibrosis in diabetic patients.

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