23,893 research outputs found
Bioactive sphene-based ceramic coatings on cpTi substrates for dental implants: An in vitro study
Titanium implant surface modifications have been widely investigated to favor the process of osseointegration. The present work aimed to evaluate the effect of sphene (CaTiSiO5) biocoating, on titanium substrates, on the in vitro osteogenic differentiation of Human Adipose-Derived Stem Cells (hADSCs). Sphene bioceramic coatings were prepared using preceramic polymers and nano-sized active fillers and deposited by spray coating. Scanning Electron Microscopy (SEM) analysis, surface roughness measurements and X-ray diffraction analysis were performed. The chemical stability of the coatings in Tris-HCl solution was investigated. In vitro studies were performed by means of proliferation test of hADSCs seeded on coated and uncoated samples after 21 days. Methyl Thiazolyl-Tetrazolium (MTT) test and immunofluorescent staining with phalloidin confirmed the in vitro biocompatibility of both substrates. In vitro osteogenic differentiation of the cells was evaluated using Alizarin Red S staining and quantification assay and real-time PCR (Polymerase Chain Reaction). When hADSCs were cultured in the presence of Osteogenic Differentiation Medium, a significantly higher accumulation of calcium deposits onto the sphene-coated surfaces than on uncoated controls was detected. Osteogenic differentiation on both samples was confirmed by PCR. The proposed coating seems to be promising for dental and orthopedic implants, in terms of composition and deposition technology
Interleukin-18 enhances vascular calcification and osteogenic differentiation of vascular smooth muscle cells through TRPM7 channel activation
ObjectiveāVascular calcification (VC) is an important predictor of cardiovascular morbidity and mortality. Osteogenic differentiation of vascular smooth muscle cells (VSMCs) is a key mechanism of VC. Recent studies show that IL-18 (interleukin-18) favors VC while TRPM7 (transient receptor potential melastatin 7) channel upregulation inhibits VC. However, the relationship between IL-18 and TRPM7 is unclear. We questioned whether IL-18 enhances VC and osteogenic differentiation of VSMCs through TRPM7 channel activation.
Approach and ResultsāCoronary artery calcification and serum IL-18 were measured in patients by computed tomographic scanning and enzyme-linked immunosorbent assay, respectively. Primary rat VSMCs calcification were induced by high inorganic phosphate and exposed to IL-18. VSMCs were also treated with TRPM7 antagonist 2-aminoethoxy-diphenylborate or TRPM7 small interfering RNA to block TRPM7 channel activity and expression. TRPM7 currents were recorded by patch-clamp. Human studies showed that serum IL-18 levels were positively associated with coronary artery calcium scores (r=0.91; P<0.001). In VSMCs, IL-18 significantly decreased expression of contractile markers Ī±-smooth muscle actin, smooth muscle 22 Ī±, and increased calcium deposition, alkaline phosphatase activity, and expression of osteogenic differentiation markers bone morphogenetic protein-2, Runx2, and osteocalcin (P<0.05). IL-18 increased TRPM7 expression through ERK1/2 signaling activation, and TRPM7 currents were augmented by IL-18 treatment. Inhibition of TRPM7 channel by 2-aminoethoxy-diphenylborate or TRPM7 small interfering RNA prevented IL-18āenhanced osteogenic differentiation and VSMCs calcification.
ConclusionsāThese findings suggest that coronary artery calcification is associated with increased IL-18 levels. IL-18 enhances VSMCs osteogenic differentiation and subsequent VC induced by Ī²-glycerophosphate via TRPM7 channel activation. Accordingly, IL-18 may contribute to VC in proinflammatory conditions
BMP2 Increases Adipogenic Differentiation in the Presence of Dexamethasone, which is Inhibited by the Treatment of TNF-alpha in Human Adipose Tissue-Derived Stromal Cells
BACKGROUND/AIMS:
The aim of this study was to analyze the effect of BMP2 on osteogenic differentiation of human adipose tissue-derived stromal cells (hADSCs).
METHODS:
Cultured cells were differentiated into osteogenic lineage in the presence of BMP2. Gene expressions were determined by real time PCR.
RESULTS:
BMP2 increased (2/8) or inhibited (6/8) osteogenic differentiation according to hADSCs batches. Regardless of the BMP2 action on osteogenic differentiation, BMP2 induced lipid droplet formation under an osteogenic differentiation condition in all batches of hADSCs, not hBMSCs, to be tested, which was confirmed by analysis of adipogenesis related genes expression. hADSCs expressed various BMP receptors. BMP2 increased expression of BMP2-responsive genes such as DLX3 and ID2, and induced SMAD1 phosphorylation in hADSCs and hBMSCs. BMP2 increased osteogenic differentiation of hADSCs in osteogenic medium in which dexamethasone was omitted. The addition of BMP2 in the control culture media containing dexamethasone alone lead to formation of lipid droplets and increased C/EBP-Ī± expression in hADSCs. In the presence of TNF-Ī±, BMP2 stimulated osteogenic differentiation of hADSCs even in hADSCs batches in which treatment of BMP2 alone inhibited osteogenic differentiation.
CONCLUSION:
These data indicate that the control of osteogenesis and adipogenesis in hADSCs is closely related, and that hADSCs have preferential commitment to adipogenic lineages
Basic fibroblastic growth factor affects the osteogenic differentiation of dental pulp stem cells in a treatmentādependent manner
AimTo determine how basic fibroblastic growth factor (bFGF) affected the osteogenic differentiation of human dental pulp stem cells (DPSCs) in vitro and in vivo.MethodologyBasic fibroblastic growth factor stimulation of DPSCs was divided into a preātreatment period and an osteogenic differentiation period. Alizarin red quantification experiments and alkaline phosphatase activity quantification assay were performed to examine the osteogenic differentiation of DPSCs after different bFGF stimulation. Quantification reverse transcription polymerase chain reaction was used to analyze the osteogenic gene expression of DPSCs after different bFGF stimulation. In addition, DPSCs that received the 1 and 2Ā weeks bFGF preātreatments as in the in vitro experiments were mineralized for 1Ā week and seeded into hydroxyapatite/tricalcium phosphate (HA/TCP) pills and subcutaneously transplanted into naked mice for 2 or 3Ā months. The transplants were removed, sliced and stained using Modified Ponceau Trichrome Stain to observe the formation of mineralized tissue.ResultsBasic fibroblastic growth factor stimulation in the osteogenic differentiation period decreased the in vitro osteogenic differentiation ability of DPSCs. One week preātreatment with bFGF increased the in vitro osteogenic differentiation ability of DPSCs, whereas 2Ā weeks preātreatment with bFGF decreased the in vitro osteogenic differentiation ability of DPSCs. The preātreatment period was vital for the osteogenic differentiation of DPSCs in vitro. The in vivo results were similar to the in vitro results.ConclusionsBasic fibroblastic growth factor affected the osteogenic differentiation of DPSCs in a treatmentādependent manner both in vitro and in vivo.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/111933/1/iej12368.pd
Activation of the Extracellular Signal-Regulated Kinase Signaling Is Critical for Human Umbilical Cord Mesenchymal Stem Cell Osteogenic Differentiation
Human umbilical cord mesenchymal stem cells (hUCMSCs) are recognized as candidate progenitor cells for bone regeneration. However, the mechanism of hUCMSC osteogenesis remains unclear. In this study, we revealed that mitogen-activated protein kinases (MAPKs) signaling is involved in hUCMSC osteogenic differentiation in vitro. Particularly, the activation of c-Jun N-terminal kinases (JNK) and p38 signaling pathways maintained a consistent level in hUCMSCs through the entire 21-day osteogenic differentiation period. At the same time, the activation of extracellular signal-regulated kinases (ERK) signaling significantly increased from day 5, peaked at day 9, and declined thereafter. Moreover, gene profiling of osteogenic markers, alkaline phosphatase (ALP) activity measurement, and alizarin red staining demonstrated that the application of U0126, a specific inhibitor for ERK activation, completely prohibited hUCMSC osteogenic differentiation. However, when U0126 was removed from the culture at day 9, ERK activation and osteogenic differentiation of hUCMSCs were partially recovered. Together, these findings demonstrate that the activation of ERK signaling is essential for hUCMSC osteogenic differentiation, which points out the significance of ERK signaling pathway to regulate the osteogenic differentiation of hUCMSCs as an alternative cell source for bone tissue engineering. Copyright Ā© 2016 Chen-Shuang Li et al
Osteogenic differentiation driven by osteoclasts and macrophages
Introduction Osteoclasts are bone-resorbing cells closely related to bone turnover, whereas different macrophage subtypes contribute to bone fracture healing. As osteoclasts and macrophages share the same hematopoietic origin, the difference between both cell types on osteoblast coupling, crosstalk extent and consequent bone formation remains poorly understood. This study compares the potential of primary cells that are routinely considered as osteoclast and macrophage cultures on their ability to support osteogenic differentiation of human mesenchymal stromal cells (hMSCs). Methods Human Peripheral Blood Mononuclear Cells (hPBMCs) were used to obtain macrophage or osteoclast cultures using appropriate stimulatory factors. With different seeding densities of hPBMCs, conditioned media from macrophage or osteoclast cultures were harvested for comparative evaluation of effects thereof on the osteogenic differentiation of hMSCs. Specific cytological staining was used to qualitatively evaluate macrophage and osteoclast cultures. Additionally, quantitative data on hMSC proliferation, osteogenic differentiation and mineralization were obtained via biochemical assays. Results Conditioned medium from osteoclast cultures obtained via low hPBMCs seeding densities, but not from high hPBMCs seeding densities or macrophages, stimulated hMSC osteogenic differentiation and mineralization. Upon cellular crosstalk, both pre-differentiated osteoclasts and non-polarized macrophages equally supported early hMSC osteogenic differentiation and mineralization, as confirmed by increased alkaline phosphatase levels within 7 days and increased calcium content within 14 days in comparison with undifferentiated controls. Initial hPBMCs seeding density strongly influences osteoclastogenesis and the paracrine effect of the resultant osteoclast population on the osteogenic differentiation of hMSCs. In addition, only in indirect coculture, macrophages provide similar stimulatory effects as pre-differentiated osteoclasts on the osteogenic differentiation of MSCs and mineralization. Conclusion Our results demonstrate stimulatory effects of osteoclast conditioned medium on hMSC osteogenic differentiation, depending on initial hPBMC seeding density. In addition, we show that osteoclast and macrophage cultures contain pools of polarized macrophages, which may be involved in the osteogenic effects. Our data provide insight into bone tissue engineering approaches by using multicellular interactions related to bone remodeling and healing for the in vitro modulation of osteogenic differentiation
\u3cem\u3eIn vitro\u3c/em\u3e Effect of Graphene Structures as an Osteoinductive Factor in Bone Tissue Engineering: A Systematic Review
Graphene and its derivatives have been wellāknown as influential factors in differentiating stem/progenitor cells toward the osteoblastic lineage. However, there have been many controversies in the literature regarding the parameters effect on bone regeneration, including graphene concentration, size, type, dimension, hydrophilicity, functionalization, and composition. This study attempts to produce a comprehensive review regarding the given parameters and their effects on stimulating cell behaviors such as proliferation, viability, attachment and osteogenic differentiation. In this study, a systematic search of MEDLINE database was conducted for in vitro studies on the use of graphene and its derivatives for bone tissue engineering from January 2000 to February 2018, organized according to the PRISMA statement. According to reviewed articles, different graphene derivative, including graphene, graphene oxide (GO) and reduced graphene oxide (RGO) with mass ratio ā¤1.5 wt % for all and concentration up to 50 Ī¼g/mL for graphene and GO, and 60 Ī¼g/mL for RGO, are considered to be safe for most cell types. However, these concentrations highly depend on the types of cells. It was discovered that graphene with lateral size less than 5 Āµm, along with GO and RGO with lateral dimension less than 1 Āµm decrease cell viability. In addition, the threeādimensional structure of graphene can promote cellācell interaction, migration and proliferation. When graphene and its derivatives are incorporated with metals, polymers, and minerals, they frequently show promoted mechanical properties and bioactivity. Last, graphene and its derivatives have been found to increase the surface roughness and porosity, which can highly enhance cell adhesion and differentiation
Impaired osteoblast differentiation in annexin A2- and -A5-deficient cells.
Annexins are a class of calcium-binding proteins with diverse functions in the regulation of lipid rafts, inflammation, fibrinolysis, transcriptional programming and ion transport. Within bone, they are well-characterized as components of mineralizing matrix vesicles, although little else is known as to their function during osteogenesis. We employed shRNA to generate annexin A2 (AnxA2)- or annexin A5 (AnxA5)-knockdown pre-osteoblasts, and determined whether proliferation or osteogenic differentiation was altered in knockdown cells, compared to pSiren (Si) controls. We report that DNA content, a marker of proliferation, was significantly reduced in both AnxA2 and AnxA5 knockdown cells. Alkaline phosphatase expression and activity were also suppressed in AnxA2- or AnxA5-knockdown after 14 days of culture. The pattern of osteogenic gene expression was altered in knockdown cells, with Col1a1 expressed more rapidly in knock-down cells, compared to pSiren. In contrast, Runx2, Ibsp, and Bglap all revealed decreased expression after 14 days of culture. In both AnxA2- and AnxA5-knockdown, interleukin-induced STAT6 signaling was markedly attenuated compared to pSiren controls. These data suggest that AnxA2 and AnxA5 can influence bone formation via regulation of osteoprogenitor proliferation, differentiation, and responsiveness to cytokines in addition to their well-studied function in matrix vesicles
Tissue inhibitor of metalloproteinase-1 (TIMP-1) regulates mesenchymal stem cells through let-7f microRNA and Wnt/Ī²-catenin signaling
Tissue inhibitor of metalloproteinases 1 (TIMP-1) is a matrix metalloproteinase (MMP)-independent regulator of growth and apoptosis in various cell types. The receptors and signaling pathways that are involved in the growth factor activities of TIMP-1, however, remain controversial. RNA interference of TIMP-1 has revealed that endogenous TIMP-1 suppresses the proliferation, metabolic activity, and osteogenic differentiation capacity of human mesenchymal stem cells (hMSCs). The knockdown of TIMP-1 in hMSCs activated the Wnt/Ī²-catenin signaling pathway as indicated by the increased stability and nuclear localization of Ī²-catenin in TIMP-1ādeficient hMSCs. Moreover, TIMP-1 knockdown cells exhibited enhanced Ī²-catenin transcriptional activity, determined by Wnt/Ī²-catenin target gene expression analysis and a luciferase-based Ī²-cateninā
activated reporter assay. An analysis of a mutant form of TIMP-1 that cannot inhibit MMP indicated that the effect of TIMP-1 on Ī²-catenin signaling is MMP independent. Furthermore, the binding of CD63 to TIMP-1 on the surface of hMSCs is essential for the TIMP-1āmediated effects on Wnt/Ī²-catenin signaling. An array analysis of microRNAs (miRNAs) and transfection studies with specific miRNA inhibitors and mimics showed that let-7f miRNA is crucial for the regulation of Ī²-catenin activity and osteogenic differentiation by TIMP-1. Let-7f was up-regulated in TIMP-1ādepleted hMSCs and demonstrably reduced axin 2, an antagonist of Ī²-catenin stability. Our results demonstrate that TIMP-1 is a direct regulator of hMSC functions and reveal a regulatory network in which let-7f modulates Wnt/Ī²-catenin activity
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Osteoprotegerin reduces osteoclast resorption activity without affecting osteogenesis on nanoparticulate mineralized collagen scaffolds.
The instructive capabilities of extracellular matrix-inspired materials for osteoprogenitor differentiation have sparked interest in understanding modulation of other cell types within the bone regenerative microenvironment. We previously demonstrated that nanoparticulate mineralized collagen glycosaminoglycan (MC-GAG) scaffolds efficiently induced osteoprogenitor differentiation and bone healing. In this work, we combined adenovirus-mediated delivery of osteoprotegerin (AdOPG), an endogenous anti-osteoclastogenic decoy receptor, in primary human mesenchymal stem cells (hMSCs) with MC-GAG to understand the role of osteoclast inactivation in augmentation of bone regeneration. Simultaneous differentiation of osteoprogenitors on MC-GAG and osteoclast progenitors resulted in bidirectional positive regulation. AdOPG expression did not affect osteogenic differentiation alone. In the presence of both cell types, AdOPG-transduced hMSCs on MC-GAG diminished osteoclast-mediated resorption in direct contact; however, osteoclast-mediated augmentation of osteogenic differentiation was unaffected. Thus, the combination of OPG with MC-GAG may represent a method for uncoupling osteogenic and osteoclastogenic differentiation to augment bone regeneration
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