-
Lasers in Medical Science Sep 2018Photobiomodulation (PBM) and photodynamic therapy (PDT) share similar mechanisms but have opposite aims. Increased levels of reactive oxygen species (ROS) in the target...
Photobiomodulation (PBM) and photodynamic therapy (PDT) share similar mechanisms but have opposite aims. Increased levels of reactive oxygen species (ROS) in the target tissue in response to light combined photosensitizer (PS) application may lead to cell proliferation or oxidative damage depending on the ROS amount. The purpose of the present study is to investigate the effect of indocyanine green (ICG)-mediated PBM on osteoblast cells by measuring cell viability, proliferation, alkaline phosphatase (ALP) activity, mineralization, and gene expressions of three phenotypic osteoblast markers. A diode laser irradiating at 809 nm (10 W output power, 50 mW/cm power density) was used at 0.5, 1, and 2 J/cm energy densities (10, 20, and 40 s respectively) was applied following ICG incubation. No inhibitory effect was observed in cell viability and proliferation according to the (4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and Alamar Blue assays. ICG-mediated PBM did not alter cell viability but increased ALP activity and enhanced mineralization of existing osteoblasts. These results were also confirmed by real-time polymerase chain reaction (RT-PCR) analysis of osteoblastic markers. PS can be combined to PBM not only to damage the malignant cells as aimed in PDT studies, but also to promote cellular activity. The findings of this in vitro study may contribute to in vivo studies and ICG-mediated PBM can have promising outcomes in bone healing and regeneration therapies in future.
Topics: Cell Line; Cell Proliferation; Cell Survival; Gene Expression Regulation; Humans; Indocyanine Green; Osteoblasts; Photochemotherapy; Photosensitizing Agents; Reactive Oxygen Species
PubMed: 29744752
DOI: 10.1007/s10103-018-2530-9 -
Biochemical and Biophysical Research... Apr 2019Discovering genes with regulatory effect of osteoblast differentiation and revealing mechanism of osteogenesis will enable us to find out more therapeutic methods for...
Discovering genes with regulatory effect of osteoblast differentiation and revealing mechanism of osteogenesis will enable us to find out more therapeutic methods for treating bone diseases. In this study, we supposed DHX58 may have a close relationship with osteogenesis and we then detected the expression of DHX58 during osteogenesis. We found that DHX58 was increased along with the osteogenic induction time extended. We then analyzed the function of DHX58 on ossification of mouse osteoblasts. The knockdown of DHX58 suppressed osteogenesis, as well as the expression of osteogenic biomarkers Runx2, OCN and Col1α1. Besides, the canonical Wnt/β-Catenin signaling pathway was found significant inhibited as DHX58 downregulated, indicating it's the downstream pathway of DHX58 in regulating osteogenesis. Furthermore, we overexpressed DHX58 and the results were in accordance with the above findings. Taken together, our results indicated that DHX58 promotes osteogenesis of mouse osteoblasts via the canonical Wnt/β-Catenin signaling pathway.
Topics: Animals; Cell Differentiation; Cell Line; Gene Knockdown Techniques; Humans; Mice; Osteoblasts; Osteogenesis; RNA Helicases; Up-Regulation; Wnt Signaling Pathway
PubMed: 30795861
DOI: 10.1016/j.bbrc.2019.02.039 -
Biochemical and Biophysical Research... Jul 2017Lamin A/C is a component of the nuclear lamina, which is involved in cellular proliferation and differentiation. However, the mechanism by which lamin A regulates...
Lamin A/C is a component of the nuclear lamina, which is involved in cellular proliferation and differentiation. However, the mechanism by which lamin A regulates osteoblast differentiation is not well understood. In this study, we investigated lamin A/C expression during osteoblast differentiation in a preosteoblastic cell line, MC3T3-E1. Real-time PCR analysis showed that lamin A/C mRNA expression was upregulated during BMP-2 induced osteoblast differentiation. Treatment with the estrogen receptor antagonist, fulvestrant, inhibited osteoblast differentiation and the upregulation of lamin A/C mRNA and protein expressions in the presence of BMP-2. These results clearly demonstrated that lamin A/C expression correlates with osteoblast differentiation. To determine the roles of lamin A expression in osteoblast differentiation, MC3T3-E1 cells were transfected with a vector overexpressing lamin A. Results showed that lamin A overexpression promoted osteoblast differentiation and calcification by inducing the expression of alkaline phosphatase, type 1 collagen, BSP, osteocalcin, and DMP-1 in the presence of BMP-2. Furthermore, lamin A overexpression partially restored osteoblastic capacity in the presence of fulvestrant by increasing the expression of BSP, osteocalcin, and DMP-1. These results suggest that lamin A plays important roles in maintaining the osteoblast differentiation and function.
Topics: Animals; Calcification, Physiologic; Cell Differentiation; Cells, Cultured; Lamin Type A; Mice; Mice, Inbred C57BL; Osteoblasts
PubMed: 28237702
DOI: 10.1016/j.bbrc.2017.02.110 -
Cells, Tissues, Organs 2009Fibroblast growth factor (FGF) signaling plays a critical role in skeletal development, yet the mechanism by which FGFs affect bone mineralization is not well...
Fibroblast growth factor (FGF) signaling plays a critical role in skeletal development, yet the mechanism by which FGFs affect bone mineralization is not well understood. Review of the literature investigating effects of FGF signaling on bone mineralization indicates that FGFs may stimulate expression of factors that prevent mineralization in the short term and enhance mineralization in the long term. Pyrophosphate is an ideal example of a factor that, dependent upon environment, has the capacity to inhibit or enhance mineralization. PC-1 is the primary generator of pyrophosphate in osteoblastic cells; therefore, regulated expression of PC-1 by FGFs may be a principal mechanism by which FGF signaling affects bone mineralization. We previously showed that FGF2 induces PC-1 expression in preosteoblastic cells and that this induction is differentiation stage dependent. In order to more directly investigate the mechanism by which PC-1 expression is regulated, we have cloned a 2.8-kb region of the PC-1 gene promoter and constructed a PC-1 gene promoter/firefly luciferase reporter construct. Results indicate that this construct is specifically responsive to FGF2 or ascorbate (an inducer of osteoblast differentiation). Promoter responsiveness to FGF2 is significantly diminished upon osteoblast differentiation, and increases in promoter activity that occur with osteoblast differentiation are inhibited by FGF2 treatment. These results indicate that the mechanism of PC-1 induction by FGF2 in preosteoblastic cells is distinct from the mechanism of induction that occurs with osteoblast differentiation. These results also indicate that PC-1 may play multiple and distinct roles in the development of mineralized tissues.
Topics: Animals; Cell Differentiation; Cell Line; Diphosphates; Fibroblast Growth Factor 2; Humans; Mice; Models, Biological; Osteoblasts; Phosphoric Diester Hydrolases; Promoter Regions, Genetic; Pyrophosphatases
PubMed: 18698132
DOI: 10.1159/000151375 -
Journal of Biomedical Materials... Oct 2009Optimization of a tissue engineering scaffold for use in bone tissue engineering requires control of many factors such as pore size, porosity, permeability and, as this...
Optimization of a tissue engineering scaffold for use in bone tissue engineering requires control of many factors such as pore size, porosity, permeability and, as this study shows, the composition of the matrix. The collagen-glycosaminoglycan (GAG) scaffold variants were fabricated by varying the collagen and GAG content of the scaffold. Scaffolds were seeded with MC3T3 osteoblasts and cultured for up to 7 days. During the culture period, osteoblastic activity was evaluated by measuring metabolic activity, cell number, and spatial distribution. Collagen and GAG concentrations both affected osteoblast viability, proliferation, and spatial distribution within the scaffold. Scaffolds containing 1% collagen (w/v) and 0.088% GAG (w/v) were found to have a porosity of approximately 99%, high cell metabolic activity and cell number, and good cell infiltration over the 7 days in culture. Taken together, these results indicate the need to tailor the parameters of a biological substrate for use in a specific tissue application, in this case bone tissue engineering.
Topics: Animals; Cell Line; Cell Proliferation; Cell Survival; Collagen; Glycosaminoglycans; Mice; Osteoblasts; Porosity; Tissue Engineering; Tissue Scaffolds
PubMed: 18767061
DOI: 10.1002/jbm.a.32207 -
Molecular and Cellular Biochemistry Oct 2007Biomechanical force is one of the major epigenetic factors that determine the form and differentiation of skeletal tissues. In this study, osteoblastic cells UMR-106...
Biomechanical force is one of the major epigenetic factors that determine the form and differentiation of skeletal tissues. In this study, osteoblastic cells UMR-106 were exposed to compressive forces at 1000 mustrain and 4000 mustrain via a four-point bending system, and analyzed by MTT and LSCM techniques. Cell proliferation activity decreased shortly after loading but recovered to normal levels within 24 h. And the cytoskeleton depolymerized at first, but then gradually repolymerized. To find out the role of cytoskeleton in mechanotransduction, we examined the relationship between cytoskeleton construction and c-fos expression. A transient stress-induced upregulation in c-fos mRNA and c-Fos protein was discovered when cells were exposed to physiological forces. And the upregulation in c-fos expression was blocked by cytochalasin D (Depolymerizing agent of microfilament). It gave clues that the organization of cytoskeleton was an important link in transcriptional control in response to low-mechanical stimulation.
Topics: Animals; Cell Proliferation; Cell Shape; Cells, Cultured; Compressive Strength; Cytoskeleton; Gene Expression Regulation; Genes, fos; Mechanotransduction, Cellular; Osteoblasts; Rats; Stress, Mechanical
PubMed: 17487456
DOI: 10.1007/s11010-007-9484-8 -
Biotechnology and Applied Biochemistry Jul 2019Osteoporosis is one of the clinical complications of long-term treatment with glucocorticoids (GCs), characterized by systemic damage of bone mass and osteoblast...
Osteoporosis is one of the clinical complications of long-term treatment with glucocorticoids (GCs), characterized by systemic damage of bone mass and osteoblast dysfunction. Hydrogen sulfide was found to be involved in GCs-induced osteoblast dysfunction. Osteoblastic MC3T3-E1 cell and mitochondrial function were determined by cell viability, M-CSF level, and ALP activity and superoxide production, membrane potential, and ATP level, respectively. The purpose of this research was to explore the impact of NaHS on osteoblastic MC3T3-E1 cell function as well as on Sirt1 and PGC1α expression in dexamethasone (DEX)-treated osteoblast cells. DEX-treated MC3T3-E1 cells exhibited decreased cell viability and ALP activity, as well as increased M-CSF level; all these changes were dramatically attenuated by NaHS. DEX-treated cells also displayed mitochondrial dysfunction, namely decreased mitochondrial membrane potential and ATP generation and increased superoxide generation, which were partly reversed by NaHS. We confirmed decreased Sirt1 and PGC1α protein expression in DEX-treated MC3T3-E1 cells by Western blot, which was also partly reversed by NaHS. Silencing of Sirt1 abrogated the protective effect of NaHS against DEX-induced cell damage and mitochondrial dysfunction. NaHS alleviates DEX-induced osteoblastic MC3T3-E1 cell injury by improving mitochondrial function.
Topics: 3T3 Cells; Animals; Cell Survival; Cells, Cultured; Dexamethasone; Mice; Mitochondria; Osteoblasts; Sulfides
PubMed: 31173404
DOI: 10.1002/bab.1786 -
Molecular Medicine Reports Aug 2017Mangiferin is a polyphenolic compound present in Salacia reticulata. It has been reported to reduce bone destruction and inhibit osteoclastic differentiation. This...
Mangiferin is a polyphenolic compound present in Salacia reticulata. It has been reported to reduce bone destruction and inhibit osteoclastic differentiation. This study aimed to determine whether mangiferin directly affects osteoblast and osteoclast proliferation and differentiation, and gene expression in MC3T3‑E1 osteoblastic cells and osteoclast‑like cells derived from primary mouse bone marrow macrophage cells. Mangiferin induced significantly greater WST‑1 activity, indicating increased cell proliferation. Mangiferin induced significantly increased alkaline phosphatase staining, indicating greater cell differentiation. Reverse transcription‑polymerase chain reaction (RT‑PCR) demonstrated that mangiferin significantly increased the mRNA level of runt‑related transcription factor 2 (RunX2), but did not affect RunX1 mRNA expression. Mangiferin significantly reduced the formation of tartrate‑resistant acid phosphatase‑positive multinuclear cells. RT‑PCR demonstrated that mangiferin significantly increased the mRNA level of estrogen receptor β (ERβ), but did not affect the expression of other osteoclast‑associated genes. Mangiferin may inhibit osteoclastic bone resorption by suppressing differentiation of osteoclasts and promoting expression of ERβ mRNA in mouse bone marrow macrophage cells. It also has potential to promote osteoblastic bone formation by promoting cell proliferation and inducing cell differentiation in preosteoblast MC3T3‑E1 cells via RunX2. Mangiferin may therefore be useful in improving bone disease outcomes.
Topics: Animals; Biomarkers; Bone Marrow Cells; Cell Differentiation; Cell Line; Cell Lineage; Cell Proliferation; Cells, Cultured; Gene Expression Profiling; Male; Mice; Osteoblasts; Osteoclasts; Xanthones
PubMed: 28627701
DOI: 10.3892/mmr.2017.6752 -
Ciba Foundation Symposium 1988The fully differentiated osteoblast may be easily recognized in bone tissue. Its cuboidal shape, its position directly opposed to the bone surface and its capacity to... (Review)
Review
The fully differentiated osteoblast may be easily recognized in bone tissue. Its cuboidal shape, its position directly opposed to the bone surface and its capacity to produce calcified bone matrix are characteristic. Three other differentiation stages are also reasonably well defined--the preosteoblast, the osteocyte and the lining cell. These differentiation stages are preceded by an unknown number of precursor, progenitor and stem cell stages. Little is known about the regulation of the transitions between the various osteogenic phenotypes and their reversibility or irreversibility. One of the reasons for this is the lack of adequate tools with which to recognize the various differentiation stages. We have developed a number of monoclonal antibodies (in bone) specifically directed against osteocytes, osteoblasts and as yet unidentified cells in the periosteum. The anti-osteocyte monoclonals were used to recognize osteocytes in bone cell cultures and we obtained purified osteocyte populations for metabolic studies. Osteocytes were shown to have binding sites for parathyroid hormone. The antibodies directed against osteoblasts showed that at present our culture conditions are inadequate to allow osteoblast differentiation in vitro.
Topics: Animals; Cell Differentiation; Humans; Osteoblasts; Osteocytes
PubMed: 3068017
DOI: 10.1002/9780470513637.ch5 -
PloS One 2017This study was designed to assess the effects of cold atmospheric plasma on osteoblastic differentiation in pre-osteoblastic MC3T3-E1 cells. Plasma was irradiated...
This study was designed to assess the effects of cold atmospheric plasma on osteoblastic differentiation in pre-osteoblastic MC3T3-E1 cells. Plasma was irradiated directly to a culture medium containing plated cells for 5 s or 10 s. Alkaline phosphatase (ALP) activity assay and alizarin red staining were applied to assess osteoblastic differentiation. The plasma-generated radicals were detected directly using an electron spin resonance-spin trapping technique. Results show that plasma irradiation under specific conditions increased ALP activity and enhanced mineralization, and demonstrated that the yield of radicals was increased in an irradiation-time-dependent manner. Appropriate plasma irradiation stimulated the osteoblastic differentiation of the cells. This process offers the potential of promoting bone regeneration.
Topics: 3T3 Cells; Alkaline Phosphatase; Animals; Cell Differentiation; Cold Temperature; Electron Spin Resonance Spectroscopy; Mice; Osteoblasts; Osteocalcin; Plasma Gases
PubMed: 28683076
DOI: 10.1371/journal.pone.0180507