Osteogenic markers are decreased in SHR rats during in vitro osteoblast differentiation

2014 ◽  
Author(s):  
Barros Thamine Landim de ◽  
Antonio Chaves-Neto ◽  
Amaral Caril do ◽  
Victor Brito ◽  
Sandra Oliveira
Keyword(s):  
Osteoblast Differentiation ◽  
Shr Rats ◽  
Osteogenic Markers

An in vitro Experimental Insight into the Osteoblast Responses to Vitamin D3 and Its Metabolites

Pharmacology ◽  
2018 ◽  
Vol 101 (5-6) ◽  
pp. 225-235 ◽  
Author(s):  
Dong Wang ◽  
Jiang Song ◽  
Huasong Ma
Keyword(s):  
Vitamin D3 ◽  
Osteoblast Differentiation ◽  
Cell Types ◽  
Cell Counting ◽  
Dependent Manner ◽  
Dihydroxyvitamin D3 ◽  
Osteogenic Markers ◽  
Polymerase Chain ◽  
Dose Dependent

Background: 25-hydroxyvitamin D3 (25[OH]VD3) has recently been found to be an active hormone. Its biological actions are also demonstrated in various cell types. However, the precise influences of vitamin D3 (VD3) and its metabolites (25[OH]VD3, 1α,25-dihydroxyvitamin D3 [1α,25-(OH)2VD3]) on the osteoblast differentiation remain largely unknown. In this work, we investigated the effects of VD3 and its metabolites in different concentrations on the early and later osteoblast differentiation and biomineralization. Methods: We first used quantitative real-time polymerase chain reaction (RT-qPCR) to evaluate the responsiveness of osteoblasts to VD3, 25(OH)VD3 or 1α,25-(OH)2VD3. We also evaluated the proliferation, differentiation and biomineralization of osteoblast at different time points via cell counting kit-8 assay and the analysis of osteogenic markers. Results: The experimental results confirmed that osteoblasts could be responsive to 25(OH)VD3 and 1α,25-(OH)2VD3 but could not directly metabolize VD3 and 25(OH)VD3. Only 200 nmol/L VD3 significantly promoted osteoblast proliferation, while 25(OH)VD3 and 1α,25-(OH)2VD3 did not show obvious actions. Moreover, the early osteogenic markers were increased by 25(OH)VD3 and 1α,25-(OH)2VD3 in a dose-dependent manner. More importantly, only 25(OH)VD3 had accelerated the gene and protein expressions of osteocalcin and the biomineralization level of osteoblasts. Conclusions: Our findings provide reliable evidence that 25(OH)VD3 at 100–200 nmol/L can induce the early and later osteoblast differentiation and biomineralization for clinical bone tissue engineering.

Anti-Osteoactivin Antibody Inhibits Osteoblast Differentiation and Function In Vitro

2003 ◽  
Vol 13 (2-4) ◽  
pp. 12 ◽  
Author(s):  
Abdulhafez A. Selim ◽  
Samir M. Abdelmagid ◽  
Reem A. Kanaan ◽  
Steven L. Smock ◽  
Thomas A. Owen ◽  
...  
Keyword(s):  
Osteoblast Differentiation ◽  
And Function

scholarly journals OP0244 A PRECLINICAL TESTING TOOL: THE IN VITRO 3D FRACTURE GAP MODEL

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 154.1-154
Author(s):  
M. Pfeiffenberger ◽  
A. Damerau ◽  
P. Hoff ◽  
A. Lang ◽  
F. Buttgereit ◽  
...  
Keyword(s):  
Fracture Healing ◽  
Bone Healing ◽  
Inflammatory Markers ◽  
Initial Phase ◽  
Gap Model ◽  
Osteogenic Markers ◽  
Immunosuppressed Patients ◽  
The Impact

Background:Approximately 10% of fractures lead to significant fracture healing disorders, with a tendency to further increase due to the aging population. Of note, especially immunosuppressed patients with ongoing inflammation show difficulties in the correct course of fracture healing leading to fracture healing disorders. Most notably, invading immune cells and secreted cytokines are considered to provide an inflammatory microenvironment within the fracture gap, primarily during the initial phase of fracture healing. Current research has the focus on small animal models, facing the problem of translation towards the human system. In order to improve the therapy of fracture healing disorders, we have developed a human cell-basedin vitromodel to mimic the initial phase of fracture healing adequately. This model will be used for the development of new therapeutic strategies.Objectives:Our aim is to develop anin vitro3D fracture gap model (FG model) which mimics thein vivosituation in order to provide a reliable preclinical test system for fracture healing disorders.Methods:To assemble our FG model, we co-cultivated coagulated peripheral blood and primary human mesenchymal stromal cells (MSCs) mimicking the fracture hematoma (FH model) together with a scaffold-free bone-like construct mimicking the bony part of the fracture gap for 48 h under hypoxic conditions (n=3), in order to reflect thein vivosituation after fracture most adequately. To analyze the impact of the bone-like construct on thein vitroFH model with regard to its osteogenic induction capacity, we cultivated the fracture gap models in either medium with or without osteogenic supplements. To analyze the impact of Deferoxamine (DFO, known to foster fracture healing) on the FG model, we further treated our FG models with either 250 µmol DFO or left them untreated. After incubation and subsequent preparation of the fracture hematomas, we evaluated gene expression of osteogenic (RUNX2,SPP1), angiogenic (VEGF,IL8), inflammatory markers (IL6,IL8) and markers for the adaptation towards hypoxia (LDHA,PGK1) as well as secretion of cytokines/chemokines using quantitative PCR and multiplex suspension assay, respectively.Results:We found via histology that both the fracture hematoma model and the bone-like construct had close contact during the incubation, allowing the cells to interact with each other through direct cell-cell contact, signal molecules or metabolites. Additionally, we could show that the bone-like constructs induced the upregulation of osteogenic markers (RUNX2, SPP1) within the FH models irrespective of the supplementation of osteogenic supplements. Furthermore, we observed an upregulation of hypoxia-related, angiogenic and osteogenic markers (RUNX2,SPP1) under the influence of DFO, and the downregulation of inflammatory markers (IL6,IL8) as compared to the untreated control. The latter was also confirmed on protein level (e.g. IL-6 and IL-8). Within the bone-like constructs, we observed an upregulation of angiogenic markers (RNA-expression ofVEGF,IL8), even more pronounced under the treatment of DFO.Conclusion:In summary, our findings demonstrate that our establishedin vitroFG model provides all osteogenic cues to induce the initial bone healing process, which could be enhanced by the fracture-healing promoting substance DFO. Therefore, we conclude that our model is indeed able to mimic correctly the human fracture gap situation and is therefore suitable to study the influence and efficacy of potential therapeutics for the treatment of bone healing disorders in immunosuppressed patients with ongoing inflammation.Disclosure of Interests:Moritz Pfeiffenberger: None declared, Alexandra Damerau: None declared, Paula Hoff: None declared, Annemarie Lang: None declared, Frank Buttgereit Grant/research support from: Amgen, BMS, Celgene, Generic Assays, GSK, Hexal, Horizon, Lilly, medac, Mundipharma, Novartis, Pfizer, Roche, and Sanofi., Timo Gaber: None declared

The effects of Pueraria thomsonii flower extract on estrogenic activity, osteoblast differentiation and osteoclast formation in vitro

Maturitas ◽  
2017 ◽  
Vol 100 ◽  
pp. 189
Author(s):  
Hosong Cho ◽  
Boyoung Lee ◽  
Wonkyung Lee ◽  
Soonran Song ◽  
Junman Lim ◽  
...  
Keyword(s):  
Osteoblast Differentiation ◽  
Estrogenic Activity ◽  
Osteoclast Formation ◽  
Flower Extract

scholarly journals Overexpression of Fibroblast Growth Factor 23 Suppresses Osteoblast Differentiation and Matrix Mineralization In Vitro

2008 ◽  
Vol 23 (6) ◽  
pp. 939-948 ◽  
Author(s):  
Hua Wang ◽  
Yuji Yoshiko ◽  
Ryoko Yamamoto ◽  
Tomoko Minamizaki ◽  
Katsuyuki Kozai ◽  
...  
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Osteoblast Differentiation ◽  
Fibroblast Growth Factor 23 ◽  
Fibroblast Growth ◽  
Matrix Mineralization

Oxysterols enhance osteoblast differentiation in vitro and bone healing in vivo

2007 ◽  
Vol 25 (11) ◽  
pp. 1488-1497 ◽  
Author(s):  
Tara L. Aghaloo ◽  
Christopher M. Amantea ◽  
Catherine M. Cowan ◽  
Jennifer A. Richardson ◽  
Ben M. Wu ◽  
...  
Keyword(s):  
Bone Healing ◽  
Osteoblast Differentiation

scholarly journals Effects of Farnesyl Pyrophosphate Accumulation on Calvarial Osteoblast Differentiation

Endocrinology ◽  
2011 ◽  
Vol 152 (8) ◽  
pp. 3113-3122 ◽  
Author(s):  
Megan M. Weivoda ◽  
Raymond J. Hohl
Keyword(s):  
Bone Formation ◽  
Osteoblast Differentiation ◽  
Biosynthetic Pathway ◽  
Squalene Synthase ◽  
Farnesyl Pyrophosphate ◽  
Matrix Mineralization ◽  
Lower Serum Cholesterol ◽  
Calvarial Osteoblast ◽  
Isoprenoid Biosynthetic Pathway

Statins, drugs commonly used to lower serum cholesterol, have been shown to stimulate osteoblast differentiation and bone formation. Statins inhibit 3-hydroxy-3-methylglutaryl (HMG)-coenzyme A reductase (HMGCR), the first step of the isoprenoid biosynthetic pathway, leading to the depletion of the isoprenoids farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP). The effects of statins on bone have previously been attributed to the depletion of GGPP, because the addition of exogenous GGPP prevented statin-stimulated osteoblast differentiation in vitro. However, in a recent report, we demonstrated that the specific depletion of GGPP did not stimulate but, in fact, inhibited osteoblast differentiation. This led us to hypothesize that isoprenoids upstream of GGPP play a role in the regulation of osteoblast differentiation. We demonstrate here that the expression of HMGCR and FPP synthase decreased during primary calvarial osteoblast differentiation, correlating with decreased FPP and GGPP levels during differentiation. Zaragozic acid (ZGA) inhibits the isoprenoid biosynthetic pathway enzyme squalene synthase, leading to an accumulation of the squalene synthase substrate FPP. ZGA treatment of calvarial osteoblasts led to a significant increase in intracellular FPP and resulted in inhibition of osteoblast differentiation as measured by osteoblastic gene expression, alkaline phosphatase activity, and matrix mineralization. Simultaneous HMGCR inhibition prevented the accumulation of FPP and restored osteoblast differentiation. In contrast, specifically inhibiting GGPPS to lower the ZGA-induced increase in GGPP did not restore osteoblast differentiation. The specificity of HMGCR inhibition to restore osteoblast differentiation of ZGA-treated cultures through the reduction in isoprenoid accumulation was confirmed with the addition of exogenous mevalonate. Similar to ZGA treatment, exogenous FPP inhibited the mineralization of primary calvarial osteoblasts. Interestingly, the effects of FPP accumulation on osteoblasts were found to be independent of protein farnesylation. Our findings are the first to demonstrate that the accumulation of FPP impairs osteoblast differentiation and suggests that the depletion of this isoprenoid may be necessary for normal and statin-induced bone formation.

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