Differentiation Of Human Osteoblast-like Cells Research Articles

Changes in the intra- and extra-mechanical environment of the nucleus in Saos-2 osteoblastic cells during bone differentiation process: Nuclear shrinkage and stiffening in cell differentiation

Osteogenic differentiation has been reportedly regulated by various mechanical stresses, including fluid shear stress and tensile and compressive loading. The promotion of osteoblastic differentiation by these mechanical stresses is accompanied by reorganization of the F-actin cytoskeleton, which is deeply involved in intracellular forces and the mechanical environment. However, there is limited information about the effect on the mechanical environment of the intracellular nucleus, such as the mechanical properties of the nucleus and intracellular forces exerted on the nucleus, which have recently been found to be directly involved in various cellular functions. Here, we investigated the changes in the intracellular force applied to the nucleus and the effect on nuclear morphology and mechanical properties during osteogenic differentiation in human osteoblast-like cells (Saos-2). We carried out cell morphological analyses with confocal fluorescence microscopy, nuclear indentation test with atomic force microscopy (AFM), and fluorescence recovery after photobleaching (FRAP) for intranuclear DNA. The results revealed that a significant reorganization of the F-actin cytoskeleton from the nuclear surfaces to the cell periphery occurred in the osteogenic differentiation processes, simultaneously with the reduction of compressive forces to the nucleus. Such changes also facilitated nuclear shrinkage and stiffening, and further intranuclear chromatin compaction. The results indicate that the reduction of the intracellular compressive force due to reorganization of the F-actin cytoskeleton affects the intra- and extra-mechanical environment of the nucleus, and this change may affect gene expression and DNA replication in the osteogenic differentiation process.

Gastrointestinal In Vitro Digests of Infant Biscuits Formulated with Bovine Milk Proteins Positively Affect In Vitro Differentiation of Human Osteoblast-Like Cells.

Infant biscuits (IBs) are part of complementary feeding from weaning up to the age of five years. They normally contain bovine milk proteins, which can influence bone development. This potential effect was investigated using experimental baked IBs, which were prepared from doughs containing different type of dairy proteins: milk protein concentrate (IB1), whey protein isolate (IB2), and skimmed milk powder (IB3). Dairy protein-free (IB0) and gluten-free (IB4) biscuits were also formulated. The in vitro gastrointestinal digests of IBs (IBDs) were tested on a co-culture of Caco-2/HT-29 70/30 cells as an in vitro model of human small intestine. None of the IBDs influenced cell viability and monolayer integrity, while IBD0 and IBD4 increased Peptide-YY production. The basolateral contents of Transwell plates seeded with Caco-2/HT-29 70/30 co-culture, mimicking metabolized IBDs (MIBDs), were tested on Saos-2 cells, an in vitro model of human osteoblast-like cells. After incubation, MIBD0, lacking dairy proteins, decreased the cell viability, while MIBD2, containing whey protein isolate, increased both the viability and the number of cells. MIBD2 and MIBD4, the latter containing both casein and whey proteins, increased alkaline phosphatase activity, a bone differentiation marker. These results highlight that IBs containing dairy proteins positively affect bone development.

Comparison of Proliferation and Differentiation of Human Osteoblast-like Cells Harvested During Implant Osteotomy Preparation Using Two Different Drilling Protocols.

Autogenous bone grafts are considered a "gold standard." The success of autografts mainly depends on their ability to promote an osteogenic response. The aim of this study was to collect autogenous bone during implant osteotomy preparation using two different drilling protocols and to evaluate and compare the proliferation and differentiation ability of the collected bone particles. Autogenous bone particles were harvested from 20 patients during implant osteotomy preparation using two different drilling protocols: (1) standard drilling protocol with saline irrigation (according to the manufacturer's recommendation) and (2) low-speed drilling protocol without saline irrigation (speed < 200 rpm). Bone samples collected were cultured in growth medium, and after 2 to 3 weeks, cells that grew out from bone grafts were cultured in the normal medium as well as in osteogenic medium for days 0, 4, 7, and 20. Scanning electron microscopy, alizarin red/toluidine blue staining, DNA, ALP, and calcium content measurements were performed. Repeated measures analysis of variance (ANOVA) with Bonferroni's test was employed to analyze the data of this study. The total DNA content was significantly higher for the low-speed drilling samples compared with the standard drilling on day 4 (P < .05), day 7 (P < .01), and day 20 (P < .001) in the normal medium and on day 7 (P < .01) and day 20 (P < .01) in the osteogenic medium. Besides, calcium measurements and mineralized matrix formation observed with alizarin red/toluidine blue staining were significantly higher for the low-speed drilling group compared with the standard drilling group. Osteogenic efficacy (differentiation and proliferation) of autogenous bone particles collected using low-speed drilling was superior compared with standard drilling samples.

Alterations to the adhesion, growth and osteogenic differentiation of human osteoblast-like cells on nanofibrous polylactide scaffolds with diamond nanoparticles

Nanofibrous scaffolds are promising materials for engineering various types of tissues, including bone tissue. The properties of nanofibers for bone tissue engineering can be improved by adding ceramic or carbon nanoparticles, including diamond nanoparticles (DNPs). In this study, poly(l-lactide) (PLLA) nanofibrous membranes reinforced with detonation DNPs (concentration from 0 to 12.28 wt%), provided by the NanoCarbon Research Institute (Japan) under the name NanoAmando®, were prepared by a needle-less electrospinning process. The biocompatibility of these DNP-reinforced PLLA membranes was tested in vitro using two human lines of osteoblast-like cells, MG-63 and Saos-2. Microscopic and biochemical methods were used. In addition, the expression of cell survival markers, focal adhesion proteins and osteogenic markers were studied in both cell types at mRNA level and at protein level. The mitochondrial activity of both cells types decreased with increasing DNP concentration in the PLLA matrix. At the same time, the expression of cyclin D and survivin, i.e. factors supporting cell growth and survival were downregulated on the DNP-loaded PLLA scaffolds, while KLF6, an antiproliferative factor, was often upregulated, at least in MG-63 cells. Thus, loading the nanofibrous polymeric scaffolds with detonation DNPs had rather adverse effects on the behavior of human bone-derived cell lines. These results are mostly in conflict with our earlier studies on poly(lactide-co-glycolide) (PLGA) scaffolds, loaded with DNPs prepared by etching the diamond layer fabricated by radiofrequency plasma-assisted chemical vapour deposition. The cell behavior on DNP-loaded scaffolds is therefore markedly influenced by the origin and the properties of the DNPs.

Incorporation of silica nanoparticles to PLGA electrospun fibers for osteogenic differentiation of human osteoblast-like cells.

The development of bone tissue engineering scaffolds still remains a challenging field, although various biomaterials have been developed for this purpose. Electrospinning is a promising approach to fabricate nanofibers with an interconnected porous structure, which can support cell adhesion, guide cell proliferation and regulate cell differentiation. The aim of this study is to fabricate composite fibers composed of poly(lactic-co-glycolic acid) (PLGA) and silica nanoparticles (NPs) via electrospinning and investigate the effect of PLGA/SiO2 composite fibers on the cellular response of osteoblast-like cells (SaOS-2 cells). SEM and EDX analysis showed that silica NPs were homogenously dispersed in the composite fibers. The mechanical behavior of the fibers showed that silica NPs acted as reinforcements at concentrations of 2.5 and 5 mg/ml. The incorporation of silica NPs led to enhancement of cell attachment and spreading on PLGA/SiO2 composite fibers. SaOS-2 cells cultured on PLGA/SiO2 composite fibers exhibited increased alkaline phosphatase activity, collagen secretion and bone nodules formation. The bone nodules formation of SaOS-2 cells increased along with the amount of incorporated silica NPs. The present findings indicate that PLGA/SiO2 composite fibers can stimulate osteogenic differentiation of SaOS-2 cells and may be a promising candidate scaffold for bone tissue engineering.

Bioactivity and mineralization of natural hydroxyapatite from cuttlefish bone and Bioglass® co-sintered bioceramics.

In this study, bioactive hydroxyapatite (HAP)-based bioceramics starting from cuttlefish bone powders have been prepared and characterized. In particular, fragmented cuttlefish bone was co-sintered with 30 wt% of Bioglass® -45S5 to synthesize HAP-based powders with enhanced mechanical properties and bioactivity. Commercial synthetic HAP was treated following the same procedure and used as a reference. The structure and composition of the bioceramics formulations were characterized using Fourier transform infrared spectroscopy, X-ray diffraction and scanning electron microscopy. After the thermal treatment of cuttlefish bone powder added with 30 wt% Bioglass, new phases with compositions of sodium calcium phosphate [Na3 Ca6 (PO4 )5 ], β-tricalcium phosphate [Ca3 (PO4 )] and amorphous silica were detected. In vitro cell culture studies were performed by evaluating proliferation, metabolic activity and differentiation of human osteoblast-like cells (MG63). Scaffolds made with cuttlefish bone powder exhibited increased apatite deposition, alkaline phosphatase activity and cell proliferation compared with commercial synthetic HAP. In addition, the ceramic compositions obtained after the combination with Bioglass® further enhanced the metabolic activity of MG63 cell and promoted the formation of a well-developed apatite layer after 7 days of incubation in Dulbecco's modified Eagle's medium.

Poly(L-lactide) nanofibrous meshes loaded with detonation diamond particles as substrates for the adhesion, growth and osteogenic differentiation of human osteoblast-like cells

Poly(L-lactide) nanofibrous meshes loaded with detonation diamond particles as substrates for the adhesion, growth and osteogenic differentiation of human osteoblast-like cells

Investigation of low-level laser therapy potentiality on proliferation and differentiation of human osteoblast-like cells in the absence/presence of osteogenic factors

Several studies have shown that low-level laser irradiation (LLLI) has beneficial effects on bone regeneration. The objective of this study was to examine the in vitro effects of LLLI on proliferation and differentiation of a human osteoblast-like cell line (Saos-2 cell line). Cultured cells were exposed to different doses of LLLI with a semiconductor diode laser (659 nm; 10 mW power output). The effects of laser on proliferation were assessed daily up to seven days of culture in cells irradiated once or for three consecutive days with laser doses of 1 or 3 J/cm(2). The obtained results showed that laser stimulation enhances the proliferation potential of Saos-2 cells without changing their telomerase pattern or morphological characteristics. The effects on cell differentiation were assessed after three consecutive laser irradiation treatments in the presence or absence of osteo-inductive factors on day 14. Enhanced secretion of proteins specific for differentiation toward bone as well as calcium deposition and alkaline phosphatase activity were observed in irradiated cells cultured in a medium not supplemented with osteogenic factors. Taken together these findings indicate that laser treatment enhances the in vitro proliferation of Saos-2 cells, and also influences their osteogenic maturation, which suggest it is a helpful application for bone tissue regeneration.

Differentiation of human osteoblast-like SaOS-2 cells

Objective To observe the course of induction of human osteoblast-like SaOS-2 cells.Methods Differentation of human osteoblast-like SaOS-2 cells was induced.The change of osteoblast phenotype was observed by alkaline phosphatase (ALP) and alizarin red staining,and the difference in the osteocalcin (OC) expression was analyzed by using realtime-PCR technique during the induction of SaOS-2 cells.Results The IA values of ALP activity on the day 0,3,6,9 of osteoblast differentiation were 163 547.0 ±847.7,236 090.0 ±967.6,28 194.9 ± 185.0,and 11665.2 ± 151.0 respectively.The IA values of ALP activity on the day 3 were significantly higher than those on the day 0,6,9 of osteoblast differentiation(P ＜ 0.05).Many mineralization nodules were seen on the day 9 of osteoblast differentiation.Realtime quantitative polymerase chain reaction revealed that the expression level of OC gene in SaOS-2 cells was significantly higher on the day 6 and 9 of osteoblast differentiation than on the day 0 and 3 (P ＜0.01).Conclusion SaOS-2 cells can be induced into mature osteoblasts. Key words: Osteoblast-like SaOS-2 cells; Osteoblast differentiation; Osteocalcin

Calcitonin gene-related peptide stimulates BMP-2 expression and the differentiation of human osteoblast-like cells in vitro

To investigate whether bone morphogenic protein-2 (BMP-2) expression was involved in calcitonin gene-related peptide (CGRP)-induced osteogenesis in human osteoblast-like cells in vitro. MG-63 osteogenic human osteosarcoma cells were treated with CGRP (10-8 mol/L) for 48 h. Cell cycle phases were determined using flow cytometry assay. The protein levels of BMP-2, ALP, Osteocalcin, ColIa1, CREB, and pCREB were measured with Western blotting, while the mRNA level of BMP-2 was measured with qR-T PCR. The expression of ALP in MG-63 cells was also studied using immunofluorescence staining. The level of cAMP was measured with ELISA assay. CGRP treatment significantly stimulated proliferation of MG-63 cells, and increased the expression of BMP-2 and the osteogenic proteins ALP, Osteocalcin and ColIa1. Pretreatment with the BMP signaling inhibitor Noggin (100 ng/mL) did not affect CGRP-stimulated proliferation and BMP-2 expression, but abolished the CGRP-induced increases of the osteogenic proteins ALP, Osteocalcin and ColIa1. Furthermore, CGRP treatment markedly increased cAMP level in MG-63 cells, whereas pretreatment with the cAMP pathway inhibitor H89 (5 μmol/L) abolished the CGRP-induced increases of cAMP level and BMP-2 expression. In MG-63 cells, the BMP pathway is involved in CGRP-induced osteogenic differentiation but not in proliferation, whereas the cAMP/pCREB pathway is involved in the expression of BMP-2.

The Effect of Surface Processing of Titanium Implants on the Behavior of Human Osteoblast-Like Saos-2 Cells

The surface qualities of dental implants appear to modulate osteoblasts' growth and differentiation, affecting bone healing. During manufacturing of implants, the surface quality is affected by industrial processes. To examine the effect of manufacturing procedures on the growth and differentiation of human osteoblast-like cells, Saos-2. Saos-2 cells were cultured on titanium (Ti) disks. Cell growth was examined using the XTT assay, and cell differentiation was tested by alkaline phosphatase (ALP) activity and osteocalcin secretion. The following variables were examined: roughening of the surface by sandblasting and acid-etching, aging of the acid used for etching, fluoride modification of the surface, and the type of the packaging material. An inverse relationship was noted between Saos-2 growth and ALP activity on the tested surfaces. Roughening of the surface tended to decrease cell proliferation and to increase differentiation. Immersion of up to 200 cycles in acid decreased proliferation and increased differentiation. Cells grown on fluoride-modified surfaces exhibited more ALP activity as compared to the unmodified surfaces. No difference was noted between the three packaging materials tested. The data suggests that industrial processes may affect the behavior of osteoblast-like cells around titanium implants and should be monitored carefully by bioassays.

Effect of lactoferrin-embedded collagen membrane on osteogenic differentiation of human osteoblast-like cells

Lactoferrin (LF) has the ability to promote the proliferation and differentiation of osteoblasts, suggesting its potential utility as an osteogenic growth factor in bone tissue engineering. However, this type of application requires improved drug delivery system (DDS) technology at the target site. In this study, we report enhanced calcium deposition and alkaline phosphatase (ALP) activity using the type I collagen membrane during osteogenic differentiation of MG63 human osteoblast-like cells, indicating that type I collagen not only acts as a site for calcification but also promotes the expression of differentiated phenotypes. We also used this membrane as a drug delivery carrier for bovine LF. Approximately 27% of LF embedded on the type I collagen membrane was released within the first hour in cell-free condition. This initial burst release of LF was followed by a slower release from the collagen membrane. Bovine LF embedded in the type I collagen membrane promoted its calcification during osteogenic differentiation of MG63 cells without the loss of LF bioactivity. Taken together, ALP activity and osteocalcin production were enhanced in the MG63 cells plated on the LF-embedded collagen membrane, suggesting that LF incorporated in the collagen membrane promoted bone-like tissue formation by MG63 cells. These observations suggest that the type I collagen membrane is useful as a drug delivery carrier for LF in bone tissue engineering.

Initial effects of low-level laser therapy on growth and differentiation of human osteoblast-like cells

Low-level laser therapy is a clinically well established tool for enhancement of wound healing. In vitro studies have also shown that low level laser therapy has a biostimulatory effect on cells of different origin. The aim of this in vitro study was to investigate the initial effect of low-level laser therapy on growth and differentiation of human osteoblast-like cells. SaOS-2 cells were irradiated with laser doses of 1 J/cm2 and 2 J/cm2 using a diode laser with 670 nm wave length and an output power of 400 mW. Untreated cells were used as controls. At 24 h, 48 h and 72 h post irradiation, cells were collected and assayed for viability of attached cells and alkaline phosphatase specific activity. In addition, mRNA expression levels of osteopontin and collagen type I were assessed using semi-quantitative RT-PCR. Over the observation period, cell viability, alkaline phosphatase activity and the expression of osteopontin and collagen type I mRNA were slightly enhanced in cells irradiated with 1 J/cm2 compared with untreated control cells. Increasing the laser dose to 2 J/cm2 reduced cell viability during the first 48 h and resulted in persistently lower alkaline phosphatase activity compared with the other two groups. The expression of osteopontin and collagen type I mRNA slightly decreased with time in untreated controls and cells irradiated with 1 J/cm2, but their expression was increased by treatment with 2 J/cm2 after 72 h. These results indicate that low-level laser therapy has a biostimulatory effect on human osteoblast-like cells during the first 72 h after irradiation. Further studies are needed to determine the potential of low-level laser therapy as new treatment concept in bone regeneration.

Type III collagen is essential for growth acceleration of human osteoblastic cells by ascorbic acid 2-phosphate, a long-acting vitamin C derivative

Collagen has been reported to be essential for the proliferation of various kinds of cells including human osteoblastic cells [Takamizawa, S., Maehata, Y., Imai, K., Senoo, H., Sato, S., Hata, R., 2004. Effects of ascorbic acid and ascorbic acid 2-phosphate, a long-acting vitamin C derivative, on the proliferation and differentiation of human osteoblast-like cells. Cell Biol. Int. 28, 255–265], but the type(s) of collagen responsible for growth regulation is not known. Presently we found that ascorbic acid 2-phosphate, a long-acting vitamin C derivative, stimulated both cell growth and the expression of mRNA for type III collagen in human osteoblast-like MG-63 cells and in normal human osteoblasts, as well as in human bone marrow mesenchymal stem cells, but not the expression of type I collagen in these cells. Epidermal growth factor also stimulated both cell growth and expression of type III collagen mRNA in MG-63 cells. Among MG-63 cell clones, their growth rates correlated significantly with their COL3A1 messenger RNA levels but not with their COL1A1 or COL1A2 messenger RNA levels. Transfection of MG-63 cells with siRNA for COL3A1 but not with that for COL1A1 decreased the growth rates of the transfected cells concomitant with a drop in the level of COL3A1 mRNA. Furthermore, cell proliferation as observed by thymidine incorporation into DNA and cell number was increased when MG-63 cells were cultured on type III collagen-coated dishes. Taken together, our results indicate that type III collagen is the collagen component responsible for the growth stimulation of human osteoblastic cells.

Cryopreservation of Osteoblast-Like Cells: Viability and Differentiation with Replacement of Fetal Bovine Serum in vitro

In reconstructive medicine, the clinical use of cryopreservation techniques depends on the absence of infectious agents such as prions. Therefore, we investigated the viability and differentiation of human osteoblast-like cells during replacement of fetal bovine serum in vitro. The aim of the present study is to replace the potentially infectious supplement fetal bovine serum during the cryopreservation procedure in order to perform future clinical trials. We used a cryopreservation technique with Me₂SO for human osteoblast-like cells of iliac cancellous bone. In the cell culture of cryopreserved and fresh osteoblast-like cells, we substituted Dulbecco’s modification of Eagle’s medium (DMEM)/Ham’s F12 plus 1% penicillin/streptomycin with autologous serum, human serum albumin and Biseko® for fetal bovine serum. For the fourth treatment group, we removed fetal bovine serum without replacing it. DMEM/Ham’s F12 plus 1% penicillin/streptomycin with fetal bovine serum served as the control group. After 4, 7, 14 and 21 days of culture for the cryopreserved and noncryopreserved cells, we performed cell counting, a WST-1 test, ELISA for collagen type I, and osteocalcin. The activity of alkaline phosphatase was also measured. The best results were obtained for the group with autologous serum as a supplement after thawing, exceeding the other groups with regard to proliferation rate. Most viable cells were observed with no replacement before freezing and after thawing of the cells. With regard to differentiation, the cultures with autologous serum after thawing of the cells showed little concentration of the differentiation markers, probably due to early contact inhibition of the cells in vitro. With regard to effort and outcome, the most promising group for cryopreservation was the one with DMEM/Ham’s F12 plus 1% penicillin/streptomycin alone before freezing, especially when osteoblast-like cells were cultured in medium with autologous serum after thawing. This is important, as this in vitro setting resembles the in vivo situation when cryopreserved bone is transplanted. These findings indicate that, for clinical purposes, fetal bovine serum can be removed for cryopreservation of iliac cancellous bone with minor loss of viability.

In Vitro Growth and Differentiation of Osteoblast-Like Cells on Hydroxyapatite Ceramic Granule Calcified From Red Algae

The purpose of this study was to analyze the interaction between osteoblast-like cells isolated from mandibular bone and hydroxyapatite ceramic bone substitute obtained from calcified red algae to assess the growth and differentiation of adherent cells on this biomaterial. The macroporous ceramic material C GRAFT/Algipore (The Clinician's Preference LLC, Golden, CO) is composed of 100% hydroxyapatite and possesses specific mechanical and physiochemical properties. Osteoblast-like cells were seeded on 200 mg of biomaterial and cultured for 6 and 21 days under osteogenic differentiation conditions. Specific alkaline phosphatase activity, DNA, and protein content of the proliferating cells were analyzed. The morphology of the cells in contact with the biomaterial was examined by scanning electron microscopy. The osteoblastic phenotype of the cells was confirmed by analysis of the expression of bone-specific genes (osteocalcin, osteopontin and collagen type I) by semi-quantitative reverse transcriptase polymerase chain reaction. The DNA and protein content increased over the culture period. Scanning electron microscopy showed cells spreading on the surface of the biomaterials, covering the macropores, and colonizing the depth of the particles. The analysis of the expression patterns of bone-related genes confirmed the osteoblastic phenotype of the cultured cells. The results of this study showed that hydroxyapatite ceramic bone substitute obtained from calcified red algae support the proliferation and differentiation of human osteoblast-like cells on its surface in vitro and might be suitable for use as scaffolds in tissue engineering strategies in vivo.

Effect of laser therapy on attachment, proliferation and differentiation of human osteoblast-like cells cultured on titanium implant material

The aim of this in vitro study was to investigate the effect of low-level laser therapy (LLLT) on the attachment, proliferation, differentiation and production of transforming growth factor-ß 1 (TGF- β 1) by human osteoblast-like cells (HOB). Cells derived from human mandibular bone were exposed to GaAlAs diode laser at dosages of 1.5 or 3 J/cm 2 and then seeded onto titanium discs. Non-irradiated cultures served as controls. After 1, 3 and 24 h, cells were stained and the attached cells were counted under a light microscope. In order to investigate the effect of LLLT on cell proliferation after 48, 72 and 96 h, cells were cultured on titanium specimens for 24 h and then exposed to laser irradiation for three consecutive days. Specific alkaline phosphatase activity and the ability of the cells to synthesize osteocalcin after 10 days were investigated using p-nitrophenylphosphate as a substrate and the ELSA-OST-NAT immunoradiometric kit, respectively. Cellular production of TGF- β 1 was measured by an enzyme-linked immunosorbent assay (ELISA), using commercially available kits. LLLT significantly enhanced cellular attachment ( P < 0.05 ). Greater cell proliferation in the irradiated groups was observed first after 96 h. Osteocalcin synthesis and TGF- β 1 production were significantly greater ( P < 0.05 ) on the samples exposed to 3 J/cm 2. However, alkaline phosphatase activity did not differ significantly among the three groups. These results showed that in response to LLLT, HOB cultured on titanium implant material had a tendency towards increased cellular attachment, proliferation, differentiation and production of TGF- β 1, indicating that in vitro LLLT can modulate the activity of cells and tissues surrounding implant material.

Effects of ascorbic acid and ascorbic acid 2-phosphate, a long-acting vitamin C derivative, on the proliferation and differentiation of human osteoblast-like cells.

In order to investigate the effect of ascorbic acid (AsA) and ascorbic acid 2-phosphate (Asc 2-P), a long-acting vitamin C derivative, on the growth and differentiation of human osteoblast-like cells, we supplemented the culture medium of MG-63 cells with various concentrations (0.25 to 1 mM) of these factors. Asc 2-P significantly stimulated nascent cell growth at all concentrations in the presence of fetal bovine serum (FBS). On the other hand, AsA showed a growth repressive effect depending on its concentration, and that of FBS. Asc 2-P also increased expression of osteoblast differentiation markers, such as collagen synthesis and alkaline phosphatase (ALP) activity. These stimulative activities of Asc 2-P were attenuated by inhibitors of collagen synthesis, indicating that these effects were dependent on collagen synthesis. Electron micrographs of the cells showed the formation of a three-dimensional tissue-like structure endowed with a mature extracellular matrix in the presence of Asc 2-P.

Three-dimensional, bioactive, biodegradable, polymer-bioactive glass composite scaffolds with improved mechanical properties support collagen synthesis and mineralization of human osteoblast-like cells in vitro.

In the past decade, tissue engineering-based bone grafting has emerged as a viable alternative to biological and synthetic grafts. The biomaterial component is a critical determinant of the ultimate success of the tissue-engineered graft. Because no single existing material possesses all the necessary properties required in an ideal bone graft, our approach has been to develop a three dimensional (3-D), porous composite of polylactide-co-glycolide (PLAGA) and 45S5 bioactive glass (BG) that is biodegradable, bioactive, and suitable as a scaffold for bone tissue engineering (PLAGA-BG composite). The objectives of this study were to examine the mechanical properties of a PLAGA-BG matrix, to evaluate the response of human osteoblast-like cells to the PLAGA-BG composite, and to evaluate the ability of the composite to form a surface calcium phosphate layer in vitro. Structural and mechanical properties of PLAGA-BG were measured, and the formation of a surface calcium phosphate layer was evaluated by surface analysis methods. The growth and differentiation of human osteoblast-like cells on PLAGA-BG were also examined. A hypothesis was that the combination of PLAGA with BG would result in a biocompatible and bioactive composite, capable of supporting osteoblast adhesion, growth and differentiation, with mechanical properties superior to PLAGA alone. The addition of bioactive glass granules to the PLAGA matrix resulted in a structure with higher compressive modulus than PLAGA alone. Moreover, the PLAGA-BA composite was found to be a bioactive material, as it formed surface calcium phosphate deposits in a simulated body fluid (SBF), and in the presence of cells and serum proteins. The composite supported osteoblast-like morphology, stained positively for alkaline phosphatase, and supported higher levels of Type I collagen synthesis than tissue culture polystyrene controls. We have successfully developed a degradable, porous, polymer bioactive glass composite possessing improved mechanical properties and osteointegrative potential compared to degradable polymers of poly(lactic acid-glycolic acid) alone. Future work will focus on the optimization of the composite scaffold for bone tissue-engineering applications and the evaluation of the 3-D composite in an in vivo model.

Orthosilicic acid stimulates collagen type 1 synthesis and osteoblastic differentiation in human osteoblast-like cells in vitro

Orthosilicic acid stimulates collagen type 1 synthesis and osteoblastic differentiation in human osteoblast-like cells in vitro