Differentiation Of SaOS-2 Cells Research Articles

Effects of Daidzein, Tempeh, and a Probiotic Digested in an Artificial Gastrointestinal Tract on Calcium Deposition in Human Osteoblast-like Saos-2 Cells.

Adequate calcium intake is crucial for the prevention and treatment of bone-related issues. Developing a nutritional source of readily bioavailable calcium is particularly significant for individuals deficient in this essential element and at risk of developing osteoporosis. This research aimed to evaluate the impact of tempeh (T), daidzein (D), and Lactobacillus acidophilus (LA) within a simulated intestinal environment consisting of Caco-2 epithelial and Saos-2 cells, focusing on their implications for bone mineralization mechanisms. In the initial phase, calcium bioaccessibility from calcium citrate (CaCt), LA, D, the daidzein combination D-CaCt-LA (D1:1:1), and the tempeh combination T-CaCt-LA (T1:1:1) was assessed through digestion simulation. The calcium content of both untreated and digested samples was determined using atomic absorption spectrometry (AAS). In the subsequent stage, the digested samples were used to induce intestinal absorption in differentiated enterocyte-like Caco-2 cells. The permeable fractions were then evaluated in a culture of osteoblast-like Saos-2 cells. Preliminary cellular experiments employed the MTT assay to assess cytotoxicity. The results indicated that the analyzed products did not influence the deposition of extracellular calcium in Saos-2 cells cultured without mineralization stimulators. The combined formulations of permeable fractions of digested CaCt, LA, D, and T demonstrated the capacity to enhance the proliferation of Saos-2 cells. In Saos-2 cells, D, D1:1:1, and LA showed no discernible impact on intracellular calcium accumulation, whereas T and T1:1:1 reduced the calcium deposits. Additionally, mRNA transcripts and alkaline phosphatase (ALP) activity levels in Saos-2 cells cultured without mineralization induction were unaffected by the analyzed products. An examination of the products revealed no discernible effect on ALP activity or mRNA expression during Saos-2 cell differentiation. Our findings suggest that tempeh, daidzein, and L. acidophilus did not positively impact cellular calcium deposition in Saos-2 cells. However, tempeh, daidzein and its combination, and L. acidophilus might enhance the process of osteogenic differentiation in Saos-2 cells. Nevertheless, this study did not identify any synergistic impact on calcium deposition and the process of osteogenic differentiation in Saos-2 cells of isoflavones and probiotics.

The Biocompatibility and Antibacterial Properties of the CNTs-Doped Magnesium Based Composite Implants in a Long-Term Biodegradation Process

The aim of this paper is to increase a new biodegradable implant material’s biodegradability, biocompatibility, and osteoinductivity in the long-term degradation process, as well as its antibacterial properties, novel carbon nanotubes (CNTs) with or without Cu element were doped into calcium phosphate (CaP)–chitosan (CS) layers and then fabricated to obtain the magnesium (Mg) matrix composites. In this paper, we investigated the influences of the CNTs-CaP-CS/Mg composites on proliferation and osteogenic differentiation of human osteosarcoma cell (SaOS-2) and human bone marrow mesenchymal stem cells (hBMSCs). Furthermore, the Cu/CNTs-CaP-CS/Mg was prepared to improve the bioactivity and antibacterial activity of the composites. The results indicated that CNTs-CaP-CS/Mg composites were suitable for proliferation and differentiation of SaOS-2 cells. Stimulated by the CNTs-CaP-CS/Mg extracts, the ALP expression of hBMSCs increased in the first 16 days and the mineralization ability of hBMSCs was highly expressed throughout the whole process which might be through the Erk1/2 signaling pathway. After CNTs-loaded Cu element, the bioactivity of the coating was satisfactory. Moreover, this new implant exhibited excellent antibacterial properties for Escherichia coli (E. coli) and Staphylococcus albus (S. albus). Collectively, these data suggest that the CNTs-CaP-CS/Mg and Cu/CNTs-CaP-CS/Mg might be potentially applied as bone implants for future clinical use.

Simultaneously stimulated osteogenesis and anti-bacteria of physically cross-linked double-network hydrogel loaded with MgO-Ag2O nanocomposites

Hydrogels, with a three-dimensional network of water-soluble polymer and water, could simulate the critical properties of extracellular matrix, which has been widely used in bone tissue engineering. However, most of conventional hydrogels for bone regeneration are fragile and have poor osteogenic activity, which restricts their applications. In this work, a novel nanoparticle-hydrogel composite consisting of physically cross-linked double-network loaded with MgO-Ag2O nanocomposites was developed by the sol-gel method. The Mg2+ released from MgO-Ag2O nanocomposites was used as an ionic cross-linking site of sodium alginate (SA), while the hydrophobic micelles in the polyacrylamide (PAAM) network is acted as another crosslinking point. The results indicated that the novel nanoparticle-hydrogel composites had good self-recovery ability and excellent mechanical properties compared with the conventional sodium alginate (SA)/polyacrylamide (PAAM) hydrogels. Additionally, it showed a slow release of Mg and Ag ions due to the dual function of the embedding effect of hydrogels and the increasing pH of the solution induced by the hydrolysis of sodium alginate. In terms of in vitro tests, the nanoparticle-hydrogel composites showed significantly stimulatory effects on the proliferation and differentiation of SaOS-2 cells. In addition, the antibacterial effects of the nanoparticle-hydrogel composites were gradually enhanced with the increase of MgO-Ag2O content.

In Vitro and In Vivo Evaluation of Carboxymethyl Cellulose Scaffolds for Bone Tissue Engineering Applications.

The present study involves the development of citric acid-cross-linked carboxymethyl cellulose (C3CA) scaffolds by a freeze-drying process. Scaffolds were fabricated at different freezing temperatures of −20, −40, or −80 °C to investigate the influence of scaffold pore size on bone regeneration. All three scaffolds were porous in structure, and the pore size was measured to be 74 ± 4, 55 ± 6, and 46 ± 5 μm for −20, −40, and −80 °C scaffolds. The pores were larger in scaffolds processed at −20 °C compared to −40 and −80 °C, indicating the reduction in pore size of the scaffolds with a decrease in freezing temperature. The cytocompatibility, cell proliferation, and differentiation in C3CA scaffolds were assessed with the Saos-2 osteoblast cell line. These scaffolds supported the proliferation and differentiation of Saos-2 cells with significant matrix mineralization in scaffolds processed at −40 °C. Subcutaneous implantation of C3CA scaffolds in the rat model was investigated for its ability of vascularization and new matrix tissue formation. The matrix formation was observed at the earliest of 14 days in the scaffolds when processed at −40 °C while it was observed only after 28 days of implantation with the scaffolds processed at −20 and −80 °C. These results suggest that the citric acid-cross-linked CMC scaffolds processed at −40 °C can be promising for bone tissue engineering application.

Powder metallurgical Ti-Mg metal-metal composites facilitate osteoconduction and osseointegration for orthopedic application

In this work, TiMg metal-metal composites (MMCs) were successfully fabricated by spark plasma sintering (SPS). In vitro, the proliferation and differentiation of SaOS-2 cells in response to TiMg metal-metal composites (MMCs) were investigated. In vivo, a rat model with femur condyle defect was employed, and TiMg MMCs implants were embedded into the femur condyles. Results showed that TiMg MMCs exhibited enhanced cytocompatibility to SaOS-2 cells than pure Ti. The micro-computed tomography (Micro-CT) results showed that the volume of bone trabecula was significantly more abundant around TiMg implants than around Ti implants, indicating that more active new-bone formed around TiMg MMCs implants. Hematoxylin-eosin (H&E) staining analysis revealed significantly greater osteointegration around TiMg implants than that around Ti implants.

The development of Cu-incorporated micro/nano-topographical bio-ceramic coatings for enhanced osteoblast response

In order to take advantages of the structural complexity of hierarchical topography and benefits of Cu, novel Cu-incorporated micro/nano-topographical coatings were developed on titanium (Ti) substrates using micro-arc oxidation (MAO) and hydrothermal treatment. A post heat treatment was employed to alter the morphology of the nanostructures and modulate the release of Cu2+. The structural evolution, phase composition, surface wettability, ion release and protein adsorption of the coatings were investigated. The results showed that the MAO-fabricated coating (denoted as MAO surface) was composed of porous TiO2 matrix distributed with CaO, SiO2 and CaSiO3 amorphous compounds. After hydrothermal treatment, a thin layer of nano-particles (20 nm in diameter) composed of CuO and calcium silicate hydrate formed on MAO surface (denoted as MAO-HT surface). These nano-particles grew to larger ones (65 nm in diameter) composed of crystallized CaO·3CuO·4TiO2 and CaO·TiO2·SiO2 after post heat treatment (denoted as MAO-HT2 surface). Moreover, the ion release rate and surface hydrophilicity of various coatings followed the trend: MAO surface > MAO-HT surface > MAO-HT2 surface. The amounts of adsorbed proteins were comparable on all surfaces. Furthermore, SaOS-2 cell response to various surfaces was investigated. The results showed that the viability of cells was significantly inhibited on MAO-HT surface due to the cyto-toxicity of high concentration Cu2+ release. However, the attachment, proliferation and differentiation of SaOS-2 cells were enhanced on MAO-HT2 surface compared to MAO surface. The results indicate that modifying Ti surface with hierarchical micro/nano-topography could lead to enhanced osteoblast response.

Boron-containing micro/nano-structured TiO2/bioceramics coatings with modulatory effects on SaOS-2 cell response

In order to take the advantages of micro/nano-topography and benefits of boron (B), B-containing micro/nano-structured TiO2/bioceramics coatings were developed on titanium (Ti) substrates for enhanced osteogenesis. The micro-arc oxidation was employed to fabricate a micro-topographical TiO2 layer (referred as Micro-Ti). Then, the second layer of nano-rods was prepared on Micro-Ti surface via hydrothermal treatment (referred as Hier-Ti). These nano-rods transformed to nano-particles after a post heat treatment (referred as Hier-HT-Ti). The adhesion, proliferation and differentiation of SaOS-2 cells were inhibited on Hier-Ti, but promoted on Hier-HT-Ti. The results indicate that the development of B-containing micro/nano-topographical coatings on Ti substrates could lead to enhanced osteogenesis.

Identification of Novel Amelogenin-Binding Proteins by Proteomics Analysis

Emdogain (enamel matrix derivative, EMD) is well recognized in periodontology. It is used in periodontal surgery to regenerate cementum, periodontal ligament, and alveolar bone. However, the precise molecular mechanisms underlying periodontal regeneration are still unclear. In this study, we investigated the proteins bound to amelogenin, which are suggested to play a pivotal role in promoting periodontal tissue regeneration. To identify new molecules that interact with amelogenin and are involved in osteoblast activation, we employed coupling affinity chromatography with proteomic analysis in fractionated SaOS-2 osteoblastic cell lysate. In SaOS-2 cells, many of the amelogenin-interacting proteins in the cytoplasm were mainly cytoskeletal proteins and several chaperone molecules of heat shock protein 70 (HSP70) family. On the other hand, the proteomic profiles of amelogenin-interacting proteins in the membrane fraction of the cell extracts were quite different from those of the cytosolic-fraction. They were mainly endoplasmic reticulum (ER)-associated proteins, with lesser quantities of mitochondrial proteins and nucleoprotein. Among the identified amelogenin-interacting proteins, we validated the biological interaction of amelogenin with glucose-regulated protein 78 (Grp78/Bip), which was identified in both cytosolic and membrane-enriched fractions. Confocal co-localization experiment strongly suggested that Grp78/Bip could be an amelogenin receptor candidate. Further biological evaluations were examined by Grp78/Bip knockdown analysis with and without amelogenin. Within the limits of the present study, the interaction of amelogenin with Grp78/Bip contributed to cell proliferation, rather than correlate with the osteogenic differentiation in SaOS-2 cells. Although the biological significance of other interactions are not yet explored, these findings suggest that the differential effects of amelogenin-derived osteoblast activation could be of potential clinical significance for understanding the cellular and molecular bases of amelogenin-induced periodontal 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

Physicochemical and cell biological characterization of PMMA bone cements modified with additives to increase bioactivity

Polymethylmethacrylate (PMMA) bone cement is the most widely used material in surgery to fix joint replacements in the bone. In this study, we propose a new approach to generate bioactive PMMA surfaces directly at the site of implantation by adding the amphiphilic molecule phosphorylated 2-hydroxyethylmethacrylate (HEMA-P) to commercial PMMA bone cement, both with or without addition of 1-5% soluble calcium and carbonate salts. The setting behavior as well as the mechanical properties, the bonding quality at the metal/cement interface, mineral deposition, and cellular response for different cement modifications were investigated in vitro. The addition of HEMA-P resulted in entirely positive effects with respect to proliferation and differentiation of osteoblast-like cells (SaOs-2) and a very tight contact at the metal/cement interface. No detrimental changes of other properties were detected. The additional incorporation of salts provoked an increased deposition of calcium phosphate minerals but no further improvement in SaOs-2 cell differentiation. A significant decrease in polarization resistance for cements with high salt content (5%) was attributed to debonding between metal and cement. The results suggest an improved clinical performance of PMMA/HEMA-P composites, which might be further enhanced by small amounts of the soluble salts.

P38 Regulates Expression of Osteoblast-specific Genes by Phosphorylation of Osterix

Osterix, a zinc finger transcription factor, is specifically expressed in osteoblasts and osteocytes of all developing bones. Because no bone formation occurs in Osx-null mice, Osterix is thought to be an essential regulator of osteoblast differentiation. We report that, in several mesenchymal and osteoblastic cell types, BMP-2 induces an increase in expression of the two isoforms of Osterix arising from two alternative promoters. We identified a consensus Sp1 sequence (GGGCGG) as Osterix binding regions in the fibromodulin and the bone sialoprotein promoters in vitro and in vivo. Furthermore, we show that Osterix is a novel substrate for p38 MAPK in vitro and in vivo and that Ser-73 and Ser-77 are the regulatory sites phosphorylated by p38. Our data also demonstrate that Osterix is able to increase recruitment of p300 and Brg1 to the promoters of its target genes fibromodulin and bone sialoprotein in vivo and that it directly associates with these cofactors through protein-protein interactions. Phosphorylation of Osterix at Ser-73/77 increased its ability to recruit p300 and SWI/SNF to either fibromodulin or bone sialoprotein promoters. We therefore propose that Osterix binds to Sp1 sequences on target gene promoters and that its phosphorylation by p38 enhances recruitment of coactivators to form transcriptionally active complexes.

1195 PSA-INDUCED OSTEOBLASTIC DIFFERENTIATION IN SAOS-2 CELLS: ROLE OF WIF-1 AND TGF-BETA

You have accessJournal of UrologyProstate Cancer: Basic Research VI1 Apr 20101195 PSA-INDUCED OSTEOBLASTIC DIFFERENTIATION IN SAOS-2 CELLS: ROLE OF WIF-1 AND TGF-BETA Nagalakshmi Nadiminty, Wei Lou, and Allen Gao Nagalakshmi NadimintyNagalakshmi Nadiminty More articles by this author , Wei LouWei Lou More articles by this author , and Allen GaoAllen Gao More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2010.02.696AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES The high prevalence of osteoblastic bone metastases in prostate cancer likely involves the production of osteoblast-stimulating factors by prostate cancer cells. The identity of the factors responsible for the homing of circulating prostate cancer cells to the bone remains unknown. Prostate specific antigen (PSA) is a serine protease produced by prostate cancer cells and is an important serological marker for prostate cancer. In this study, we examined the involvement of Wnt inhibitory factor 1 (Wif-1) and Transforming growth factor beta (TGF-beta) in osteoblastic differentiation induced by PSA in SaOS-2 human osteosarcoma cells. METHODS Microarray analysis of RNAs from Saos-2 cells stably expressing PSA and vector controls was performed and the results were validated by northern blotting. The potential role of Wif-1 and TGF-β in osteoblatic differentiation induced by PSA was examined by alkaline phosphatase assays, luciferase assays and northern analysis. RESULTS PSA was found to induce bone remodeling in SaOS-2 cells as evidenced by increases in alkaline phosphatase activity, mineral deposition and osteogenic gene expression. Saos-2 cells expressing PSA exhibited markedly upregulated expression of markers of osteoblastic differentiation like collagen type 1, osteocalcin, Runx-2, alkaline phosphatase and bone sialoprotein compared to parental Saos-2 cells and vector controls. Treatment of SaOS cells with exogenous enzymatically active PSA produced similar results, suggesting that osteoblastic differentiation may be regulated by PSA. Among the genes found to be regulated by PSA expression, TGF-beta and Wif-1 were examined further. PSA enhanced the expression and activation of TGF-beta as shown by higher levels of TGF-beta-responsive reporter activity and increase in expression of TGF-beta receptors I and II. Overexpression of Wif-1 in SaOS-2 cells enhanced osteogenic gene expression, while inhibition of Wif-1 expression in PSA-expressing cells resulted in reduction of alkaline phosphatase activity, a marker of osteoblastic differentiation. CONCLUSIONS PSA-induced bone remodeling was mediated by enhanced expression of several markers of osteoblastic differentiation and modulation of TGF-beta and the Wnt/beta-catenin pathways. These results suggest that PSA produced by metastatic prostate cancer cells may participate in bone remodeling in favor of development of osteoblastic metastases. These findings provide a molecular basis for understanding the high prevalence of osteoblastic bone metastases in prostate cancer. Sacramento, CA© 2010 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 183Issue 4SApril 2010Page: e463 Advertisement Copyright & Permissions© 2010 by American Urological Association Education and Research, Inc.MetricsAuthor Information Nagalakshmi Nadiminty More articles by this author Wei Lou More articles by this author Allen Gao More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Effects of substrates nanopatterning on osteosarcoma cells behaviour

Engineering of the cellular microenvironment has become an attractive strategy to guide cellular activities such as spreading, motility, proliferation and differentiation. From a technological perspective, the physical crosstalk between the cell and its surroundings represents a design parameter that may be modulated to achieve desired physiological outcome. In this study we present a surface engineering approach to tap into the physical crosstalk between the cell and its surroundings in order to modulate osteogenic anchorage-dependent differentiation and bone formation. The effectiveness of this approach was studied by comparing the cellular behaviour of human SOAS sarcoma cells on nanostructured silicon substrates with distinct nanoscale patterns. Random nano-islands were realized by controlled deposition of tin on the polished side of silicon wafers by thermal evaporation. Four different shaped surfaces of nano structured substrates were used in this study. Silicon substrates present surface islands with diameters ranging from 10 to 35 nm and inter-island distances of 41 (B), 51 (E) or 80 (F) nm respectively. Substrate A is planar silicon used as control. Cells were seeded at 5000/cm2 on plastic or different silicon chips. Firstly we assayed the surfaces’ adhesiveness and tested the distribution of cytoskeleton and focal adhesions molecules, no particular differences in the distribution of the different proteins were observed. Terminal differentiation of Saos-2 cells was analyzed by real time PCR. The expression of OP and BOSP genes seems to be differently regulated by the surfaces, in particular substrate E seem to exert in long term experiments the optimal response. Nanostructured substrates were observed to be superior to planar controls in the modulation of cell proliferation and differentiation. Our findings suggest that physical nano-scale topography presents an instructive background to guide cell behaviour by dictating the quality of cell adhesion and focal adhesion spatial distribution which in turn can control the fate of cells.

Effects of Low-Amplitude, High-Frequency Vibrations on Proliferation and Differentiation of SAOS-2 Human Osteogenic Cell Line

The aim of the work was to understand the consequences of low-amplitude, high-frequency vibrations on proliferation and differentiation of SAOS-2 cells (sarcoma osteogenetic), an osteoblastic and tumorigenic cell line. We realized a bioreactor composed of an eccentric motor that produces a displacement of 11 mm at frequencies between 1 and 120 Hz on a plate connected to the motor. The cultures of SAOS-2 cells were fixed on the plate, and the linear acceleration provoked by the motor to the cultures was measured. We used 30 Hz as stimulating frequency after a preliminary test on the effect of different frequencies on differentiation of cells. Afterward, SAOS-2 cells were stimulated with 30 Hz for different durations, every day for 4 days. The expression of some genes involved in the differentiation process was analyzed first with a reverse transcriptase-polymerase chain reaction and afterward with a real-time polymerase chain reaction on the most expressed genes. Moreover, the proliferation of cells was evaluated. The results suggest a strong increase in the expression of the genes involved in tissue differentiation in the treated groups with respect to the controls. On the other hand, the proliferation seems to be slowed down, so probably the acceleration perceived by the mechanosensors of the cells changes the cellular cycle by blocking the duplication to early differentiate toward bone tissue.

Modulation of osteosarcoma cell growth and differentiation by silane‐modified surfaces

The effects of growing the Saos-2 human osteosarcoma cell line onto surfaces containing -CH3, -OH, -COOH, -NH2, and C6H5 groups obtained by silane modification were examined. These cells were used because of the great importance of bone cells in many aspects of biomaterials research. Silane-modified surfaces were characterized by contact angle measurements and, subsequently, surface energies were calculated. Cells grown on clean glass, as well as those grown on glass surfaces containing the functional groups cited above, were examined by light and scanning electron microscopy and assessed for their growth characteristics (i.e., determination of cell number and Ki67 antigen expression). The data presented seemed to indicate that if Saos-2 cells are grown on silane-modifed surfaces containing the methyl (CH3), hydroxyl (OH), and phenyl (C6H5) functional groups, their proliferation is slowed down while growth of these cells on glass surfaces modified with amino (NH2) and carboxyl (COOH) groups did not significantly affect growth. Once it was demonstrated that these three functional groups induce significant variations in proliferation, cells grown on these surfaces were also tested for apoptosis and expression of important markers of bone cell differentiation (i.e., osteonectin and osteopontin) by flow cytometry and eventual rearrangement of these markers by fluorescence microscopy. The data suggested that growth of Saos-2 cells on CH3 induces the most evident morphological changes while growth of these cells on OH and C6H5 brings about the greater variations in osteonectin and osteopontin. We hypothesized that these changes are indicative of an increase in differentiation of Saos-2 cells when grown on the OH and C6H5 groups. © 2001 John Wiley & Sons, Inc. J Biomed Mater Res 55: 338–349, 2001

Modulation of osteosarcoma cell growth and differentiation by silane-modified surfaces.

The effects of growing the Saos-2 human osteosarcoma cell line onto surfaces containing -CH(3), -OH, -COOH, -NH(2), and C6H5 groups obtained by silane modification were examined. These cells were used because of the great importance of bone cells in many aspects of biomaterials research. Silane-modified surfaces were characterized by contact angle measurements and, subsequently, surface energies were calculated. Cells grown on clean glass, as well as those grown on glass surfaces containing the functional groups cited above, were examined by light and scanning electron microscopy and assessed for their growth characteristics (i.e., determination of cell number and Ki67 antigen expression). The data presented seemed to indicate that if Saos-2 cells are grown on silane-modifed surfaces containing the methyl (CH(3)), hydroxyl (OH), and phenyl (C6H5) functional groups, their proliferation is slowed down while growth of these cells on glass surfaces modified with amino (NH(2)) and carboxyl (COOH) groups did not significantly affect growth. Once it was demonstrated that these three functional groups induce significant variations in proliferation, cells grown on these surfaces were also tested for apoptosis and expression of important markers of bone cell differentiation (i.e., osteonectin and osteopontin) by flow cytometry and eventual rearrangement of these markers by fluorescence microscopy. The data suggested that growth of Saos-2 cells on CH(3) induces the most evident morphological changes while growth of these cells on OH and C6H5 brings about the greater variations in osteonectin and osteopontin. We hypothesized that these changes are indicative of an increase in differentiation of Saos-2 cells when grown on the OH and C6H5 groups.