Human Osteoblast Cell Line hFOB Research Articles

Characterization of chondroitin sulfate from deer tip antler and osteogenic properties

Deer antler is a highly regenerative tissue that involves cellular differentiation, osteogenesis and ossification processes. Chondroitin sulfate is the major glycosaminoglycan contained in antler connective tissue and has been isolated from cartilaginous antler by 4M GuHCl extraction, gradient ultracentrifugation and chromatography techniques. We examined the disaccharide composition by 2-AB labeling and anion exchange HPLC analysis of the three resultant fractions (high, medium and low density fractions). The high density fraction consists of A-unit and D-unit disaccharide in the ratio of 1:1, whereas, the CS disaccharide composition ratio of A- unit:C-unit:D-Unit:E-unit contained in medium and low density fractions are 3:4:3:1 and 2:2:2:1, respectively. The only intact CS oligosaccharides of the medium density fraction upregulated gene expression of bone-specific proteins of a human osteoblastic cell line (hFOB1.19). Thus, CS oligosaccharides from cartilaginous deer antler, with their oversulfated chondroitin sulfate composition, demonstrated the physiological properties and may be good candidates for osteogenetic agents in humans.

Plasma membrane proteomic analysis of human osteosarcoma and osteoblastic cells: revealing NDRG1 as a marker for osteosarcoma

Osteosarcoma (OS) is the most common primary malignant bone tumor in children and adolescents. To identify new biomarkers for early diagnosis of OS and novel therapeutic candidates, we carried out a plasma membrane proteomic study based on two-dimensional electrophoresis (2DE). The OS cell line MG-63 and the human osteoblastic cell line hFOB1.19 were adopted as the comparison model. We extracted plasma membrane by aqueous two-phase partition extraction. The proteins were separated through 2DE. We analyzed the differentially expressed proteins by Imagemaster software and then identified them by liquid chromatography-tandem mass spectrometry, and the location and function of differential proteins were searched through the Gene Ontology database. In total, 220 protein spots were separated by 2DE. Seven proteins with more than 2.0-folds of difference were successfully identified from 13 gel spots, with 6 up-regulated and 1 down-regulated. Gene Ontology analysis of the differentially expressed proteins indicated that these proteins were involved in seven kinds of functions including binding, structural, cell motility, receptor activity, electron carrier activity, NADH dehydrogenase (ubiquinone) activity, and transcription repressor activity. The up-regulation of NDRG1 was verified in osteosarcoma through Western blotting and by immunohistochemistry in paraffin-embedded tissues. The plasma membrane proteins identified in this study may provide new insights into osteosarcoma cancer biology and potential diagnostic and therapeutic biomarkers.

Role of the EGFR ligand/receptor system in the secretion of angiogenic factors in mesenchymal stem cells

Increasing evidence suggests that bone marrow-derived mesenchymal stem cells (MSCs) are recruited into the stroma of developing tumors where they contribute to cancer progression. MSCs produce different growth factors that sustain tumor-associated neo-angiogenesis. Since the majority of carcinomas secrete ligands of the epidermal growth factor receptor (EGFR), we assessed the role of EGFR signaling in regulating the release of angiogenic factors in MSCs. Treatment of human primary MSCs and of the human osteoblastic cell line hFOB with transforming growth factor α (TGF-α), one of the main ligands of the EGFR, significantly induced activation of this receptor and of different intracellular signaling proteins, including the PI3K/AKT and the MEK/MAPK pathways. TGF-α induced a significant increase in the levels of secretion of vascular endothelial growth factor in both MSCs and hFOB. Conditioned medium from TGF-α treated MSCs showed an higher in vivo angiogenic effect as compared with medium from untreated cells. Treatment of MSCs with TGF-α also produced a significant increase in the secretion of other angiogenic growth factors such as angiopoietin-2, granulocyte-colony stimulating factor, hepatocyte growth factor, interleukin (IL)-6, IL-8, and platelet-derived growth factor-BB. Using selective MEK and PI3K inhibitors, we found that both MEK/MAPK and the PI3K/AKT signaling pathways mediate the ability of TGF-α to induce secretion of angiogenic factors in MSCs. Finally, stimulation with TGF-α increased the ability of MSCs to induce migration of MCF-7 breast cancer cells. These data suggest that EGFR signaling regulates the ability of MSCs to sustain cancer progression through the release of growth factors that promote neo-angiogenesis and tumor cell migration.

Granulocyte-Macrophage Colony-Stimulating Factor- and Tumor Necrosis Factor Alpha-Mediated Matrix Metalloproteinase Production by Human Osteoblasts and Monocytes after Infection withBrucella abortus

Osteoarticular complications are common in human brucellosis, but the pathogenic mechanisms involved are largely unknown. Since matrix metalloproteinases (MMPs) are involved in joint and bone damage in inflammatory and infectious diseases, we investigated the production of MMPs by human osteoblasts and monocytes, either upon Brucella abortus infection or upon reciprocal stimulation with factors produced by each infected cell type. B. abortus infection of the normal human osteoblastic cell line hFOB 1.19 triggered a significant release of MMP-2, which was mediated in part by granulocyte-macrophage colony-stimulating factor (GM-CSF) acting on these same cells. Supernatants from infected osteoblasts exhibited increased levels of monocyte chemoattractant protein 1 and induced the migration of human monocytes (THP-1 cell line). Infection with B. abortus induced a high MMP-9 secretion in monocytes, which was also induced by heat-killed B. abortus and by the Omp19 lipoprotein from B. abortus. These effects were mediated by Toll-like receptor 2 and by the action of tumor necrosis factor alpha (TNF-α) produced by these same cells. Supernatants from B. abortus-infected monocytes induced MMP-2 secretion in uninfected osteoblasts, and this effect was mediated by TNF-α. Similarly, supernatants from infected osteoblasts induced MMP-9 secretion in uninfected monocytes. This effect was mediated by GM-CSF, which induced TNF-α production by monocytes, which in turn induced MMP-9 in these cells. These results suggest that MMPs could be potentially involved in the tissue damage observed in osteoarticular brucellosis.

Comparison of Tantalum and Hydroxyapatite Coatings on Titanium for Applications in Load Bearing Implants

AbstractOver the years hydroxyapatite (HA) coatings have been used to improve biologic properties of Ti‐based load bearing metal implants. However, applicability of HA coated implants is subjected to physical stability of the HA phase and mechanical integrity of the coating‐substrate interface. In this study, we have compared the microstructure and in vitro cell–materials interactions of newly developed laser deposited Ta coatings and radio frequency (RF) induction plasma sprayed HA coatings on Ti substrate. In vitro biocompatibility study, using human osteoblast cell line hFOB, showed equally excellent cellular adherence and growth on Ta and HA coatings. Quantitative assay of cell survivability on these coatings showed that the Ta coatings provide comparable initial cell attachment to that of HA coatings. Microstructural analysis of the coatings showed strong metallurgical bonding without sharp interface between the Ta coating and the Ti substrate, while the interface between HA coating and the Ti substrate was sharp. The interface microstructural features and in vitro cell–materials interactions of Ta coatings on Ti clearly demonstrate their potential to replace HA based coatings for enhanced/early biologic fixation. Other significant benefits of these dense Ta coatings include high toughness, strong bonding with the substrate, and long‐term stability of the interface.

The osteogenic differentiation of human osteoprogenitor cells on Anodic-Plasma-Chemical treated Ti6Al7Nb

Biological integration of an implant to surrounding bone is an important event for its clinical success and is driven by numerous factors, including the attraction of bone forming cells. The implant’s surface properties influence the initial cell response at the cell/material interface, ultimately affecting the rate and quality of new tissue formation and the stability of the implant. As a consequence, various surface treatments have been developed to increase the clinical performance of titanium-based implants. Among them, the Anodic Plasma-Chemical (APC) technique allows for the combined chemical and morphological modification of titanium surfaces in a single process step. In the present study, we compared the potential of APC surface treatment of high-strength titanium alloys with vacuum plasma spray treatment and yellow gold anodization in supporting osteogenic differentiation of two different osteoprogenitor cell types. Both human fetal osteoblast cell line (hFOB1.19) and human mesenchymal stromal cells showed extensive cell spreading, faster cell growth and differentiation on APC surfaces compared to vacuum plasma spray treated and yellow gold anodized surfaces. Our findings showed that APC titanium-based surfaces provided an effective substrate for osteoprogenitor cells adhesion, proliferation and differentiation.

Modifications of a calcium phosphate cement with biomolecules—Influence on nanostructure, material, and biological properties

Calcium phosphate cements (CPC), forming hydroxyapatite during the setting reaction, are characterized by good biocompatibility and osteoconductivity, however, their remodeling into native bone tissue is slow. One strategy to improve remodeling and bone regeneration is the directed modification of their nanostructure. In this study, a CPC was set in the presence of cocarboxylase, glucuronic acid, tartaric acid, α-glucose-1-phosphate, L-arginine, L-aspartic acid, and L-lysine, respectively, with the aim to influence formation and growth of hydroxyapatite crystals through the functional groups of these biomolecules. Except for glucuronic acid, all these modifications resulted in the formation of smaller and more agglomerated hydroxyapatite particles which had a positive impact on the biological performance indicated by first experiments with the human osteoblast cell line hFOB 1.19. Moreover, adhesion, proliferation, and osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSC) as well as binding of the growth factors BMP-2 and VEGF was investigated on CPC modified with cocarboxylase, arginine, and aspartic acid. Initial adhesion of hBMSC was improved on these three modifications and proliferation was enhanced on CPC modified with cocarboxylase and arginine whereas osteogenic differentiation remained unaffected. Modification of the CPC with arginine and aspartic acid, but not with cocarboxylase, led to a higher BMP-2 binding.

In vitro biocompatibility and corrosion resistance of a new implant titanium base alloy

One objective of this work was to study the corrosion resistance of the new implant Ti-10Zr-5Ta-5Nb alloy in physiological fluids of different pH values, simulating the extreme functional conditions. Another objective was in vitro biocompatibility evaluation of the new alloy using human fetal osteoblast cell line hFOB 1.19. Cytocompatibility was assessed by determination of possible material cytotoxic effects, cell morphology and cell adhesion. The thermo-mechanical processing of the new implant alloy consisted in plastic deformation (almost 90%) performed by hot rolling accompanied by an initial and final heat treatment. The new Ti-10Zr-5Ta-5Nb alloy presented self-passivation, with a large passive potential range and low passive current densities, namely, a very good anticorrosive resistance in Ringer solution of acid, neutral and alkaline pH values. Cell viability was not affected by the alloy substrate presence and a very good compatibility was noticed.

Myeloma cells inhibit osteogenic differentiation of mesenchymal stem cells and kill osteoblasts via TRAIL-induced apoptosis

IntroductionMyeloma bone disease (MBD) is the result of the increased activity of osteoclasts (OCs), which is not accompanied by a comparable increase of osteoblast (OB) function, thus leading to enhanced bone resorption. Osteoblasts can also regulate osteoclast activity through expression of cytokines, such as receptor activator of nuclear factor-κB ligand (RANKL), which activates osteoclast differentiation, and osteoprotegerin (OPG), which inhibits RANKL by acting as a decoy receptor.Material and methodsBased on a series of 21 patients with multiple myeloma (MM) and human osteoblast cell line HFOB1.19, we provide evidence that the bone marrow-derived mesenchymal stem cells (BMMSCs) of patients with MM exhibit normal phenotype, but showed reduced efficiency to differentiate into OBs as compared with normal controls.ResultsIn vitro assays showed that MM cells inhibited the potential of osteogenic differentiation of BMMSCs from healthy controls and rendered the OBs sensitive to TRAIL-induced apoptosis. There was no evidence of the formation of tartrate-resistant acid phosphatase positive OCs. The osteogenic differentiation of HFOB1.19 was also inhibited in the presence of RPMI 8266 or XG7 MM cells, as confirmed by von Kossa and ALP staining. Osteoblast s induced from BMMSCs supported survival and proliferation of MM cells, especially when the MM cells were cultured in medium containing rhTRAIL and dexamethasone. Multiple myeloma cells proliferated and grew well in the presence of residual OBs.ConclusionsBesides OCs, our results demonstrated that OBs and MM cells were dependent upon each other and made a microenvironment suitable for MM cells.

Direct laser processing of a tantalum coating on titanium for bone replacement structures

Recently tantalum is gaining more attention as a new metallic biomaterial as it has been shown to be bioactive and biologically bonds to bone. However, the relatively high cost of manufacture and an inability to produce a modular all Ta implant has limited its widespread acceptance. In this study we have successfully deposited a Ta coating on Ti using laser engineered net shaping (LENS™) to enhance the osseointegration properties. In vitro biocompatibility study, using human osteoblast cell line hFOB, showed excellent cellular adherence and growth with abundant extracellular matrix formation on the Ta coating surface compared with the Ti surface. A six times higher living cell density was observed on the Ta coating than on the Ti control surface by MMT assay. A high surface energy and wettability of the Ta surface were observed to contribute to its significantly better cell–material interactions. Also, these dense Ta coatings do not suffer from low fatigue resistance due to the absence of porosity and a sharp interface between the coating and the substrate, which is a major concern for porous coatings used for enhanced/early biological fixation.

Serum‐mediated osteogenic effect in traumatic brain‐injured patients

Patients with a traumatic brain injury (TBI) and bone fractures often show an enhanced fracture healing, as well as an increased incidence of heterotopic ossifications (HO). It has been suggested that unknown osteoinductive factors may be released by the injured brain into the systemic blood circulation and act peripherally on the affected tissues. The aim of this study was to investigate whether serum from TBI patients is osteoinductive. Sixty-one consecutive patients were classified into four groups: TBI and long-bone fracture (group I, n = 12), isolated severe TBI (group II, n = 21), isolated long-bone fracture (group III, n = 19) and controls (group IV, n = 9). Blood samples were collected at 6, 24, 72 and 168 h post-injury. The osteogenic potential was determined by measuring the in vitro proliferation rate of the human fetal osteoblastic cell line hFOB1.19, and primary human osteoblasts. Additionally, serum induced osteoblastic differentiation was assessed by measuring the mRNA expression of specific osteoblastic markers, including alkaline phosphatase, runt-related transcription factor 2, cathepsin K and serine protease 7. The sera of group I induced a higher mean proliferation rate of primary human osteoblasts at all time points of sampling than group III (P < 0.05). Group I had a higher mean proliferation rate of hFOB1.19 cells than all other groups at 6, 24 and 72 h post-injury (P < 0.05). The expression of alkaline phosphatase, cathepsin K and runt-related transcription factor 2 mRNA was increased in group I compared with group III and serine protease 7 was exclusively expressed in group I. The study results strongly support a humoral mechanism in enhanced fracture healing and the induction of HO after TBI. Increased proliferation of osteoblastic cells and an accelerated differentiation of osteoprogenitor cells may be responsible for increased osteogenesis in TBI.

Humoral Factors Enhance Fracture-Healing and Callus Formation in Patients with Traumatic Brain Injury

Scientific evidence is mounting for an association between traumatic brain injury and enhanced osteogenesis. The aim of this study was to correlate the in vitro osteoinductive potential of serum with the features of fracture-healing and the extent of brain damage in patients with severe traumatic brain injury and bone fracture. Patients with a long-bone fracture and a traumatic brain injury (seventeen patients) or without a brain injury (twenty-four patients) were recruited. The Glasgow Coma Scale score was determined on admission. Radiographs of the fracture were made before surgery, at six weeks, and at three, six, and twelve months after surgery. The time to union was estimated clinically and radiographically, and the callus ratio to shaft diameter was calculated. Serum samples were collected at six, twenty-four, seventy-two, and 168 hours after injury, and their osteogenic potential was determined by measurement of the in vitro proliferation rate of the human fetal osteoblastic cell line hFOB1.19. Patients with a traumatic brain injury had a twofold shorter time to union (p = 0.01), a 37% to 50% increased callus ratio (p < 0.01), and their sera induced a higher proliferation rate in hFOB cells (p < 0.05). A linear relationship was revealed between hFOB cell proliferation rates and the amount of callus formed (p < 0.05). The Glasgow Coma Scale score was correlated with the callus ratio on both radiographic projections (p < 0.05), time to union (p = 0.04), and the proliferation rate of hFOB cells at six hours after injury (p = 0.03). Patients with a severe brain injury release unknown humoral factors into the blood circulation that enhance and accelerate fracture-healing.

Strontium modified biocements with zero order release kinetics

Strontium-substituted β-TCP with the general formula Ca (3− x) Sr x (PO 4) 2 (0 < x < 1) was synthesized by calcination of powder mixtures with the appropriate stoichiometry and reacted with acidic monocalcium phosphate monohydrate (MCPM) to form a cementitious matrix of secondary calcium phosphates. The use of Sr-substituted β-TCP as a reactant not only induced strontium substitution in the setting products but surprisingly it also favoured the formation of monetite (CaHPO 4) as setting product, whereas Sr-free cements set to brushite (CaHPO 4·2H 2O). Release experiments under dynamic conditions for up to 15 days revealed the release of Sr 2+ ions at dose ranges of 12–30 ppm with zero order release kinetic. Cement cytocompatibility was investigated by culturing human osteoblast cell line hFOB1.19 on cement surfaces whereas Sr-containing cements were as good as Sr-free cements in providing a template for cell growth and function.

Ag and Ag/N 2 plasma modification of polyethylene for the enhancement of antibacterial properties and cell growth/proliferation

Polyethylene (PE) is one of the most common materials used for medical implants. However, it usually possesses low biocompatibility and insufficient antibacterial properties. In the work described here, plasma immersion ion implantation (PIII) is employed to implant silver into PE to enhance both its antibacterial properties and its biocompatibility. Our results show that Ag PIII can give rise to excellent antibacterial properties and induces the formation of functional groups such as C–O and C C. These C–O and C C groups on the modified surface can trigger the growth of the human fetal osteoblastic cell line (hFOB). Furthermore, combining N 2 and Ag PIII prolongs the antibacterial effects, but nitrogen-containing functional groups such as C–N and C N created by N 2 co-PIII negatively impact proliferation of hFOB on the surface. According to our experimental investigation on cell proliferation, functional groups such as C–N and C N created by nitrogen PIII are disadvantageous to cell growth whereas the C–O and C C groups benefit cell growth. Both the antibacterial activity and biocompatibility of PE can be enhanced by means of the proper plasma surface treatment.

NF-kappa B Plays a Key Role in the Hematopoietic Niche Function of Osteoblast after Ionizing Radiation.

Recent studies demonstrated that osteoblasts in the endosteal region of bone are a key cellular component of the hematopoietic niche that directly regulates hematopoietic stem cell (HSC) survival, proliferation and differentiation. In experiments with the human fetal osteoblast cell line hFOB1.19, we found survival of gamma-irradiated primary human hematopoietic CD34+ cells was significantly enhanced by co-culture with hFOB cells or by conditioned medium (CM) from hFOB cells, as shown by increased colony efficiency and colony size. In clonogenic assays of CD34+ cells starting with 5 × 103 cells/dish, co-culture with hFOB cells increased colony number from 428±53 (control) to 988±65 (co-culture) after 2 Gy (p<0.01), 301±44 to 739±56 after 4 Gy (p<0.01), and 45±15 to 72±10 after 6 Gy (p<0.05). Surprisingly, although radiation induces osteoblast cell damage, CM from 6 Gy-irradiated hFOB cells had a greater effect than CM from unirradiated hFOB cells in supporting CD34+ clonogenicity and resulted in colony number increase from 430±40 (CM from unirradiated hFOB cells) to 773±48 (CM from 6 Gy-irradiated hFOB cells) after 2 Gy IR (p<0.01). Using a cytokine array for 120 cytokines and chemokines, we detected 11 factors released from hFOB cells, and 4 of them (IL-6, G-CSF, GRO, and IL-8) were enhanced by ionizing radiation (IR). IR (2–8 Gy) caused hFOB cell cycle arrest in G2 phase after 24 h, and irreversible cell cycle block and apoptotic cell death were observed 72 h after IR in a radiation dose-dependent manner. IR induced NF-kappaB (NFkB) p65 phosphorylation and NFkB activity in hFOB cells. Inhibition of NFkB expression with siRNA blocked the ability of hFOB CM to support clonogenic survival of CD34+ cells, and abrogated the effect of IR on release of IL-6, G-CSF, GRO, and IL-8. Radiation upregulated p53 (ser15) phosphorylation within 4 h, and p21 expression after 24 h in hFOB cells. However, NFkB gene silencing accelerated p21 expression and induced a massive apoptosis of hFOB cells. We propose that IR induces NFkB and p53 activation simultaneously in hFOB cells. The fate of osteoblasts after irradiation depends on the amount and severity of injury, and on repair processes regulated by NFkB and p53. Silencing the NFkB gene disrupted this balance and resulted in hFOB cell apoptosis.

The Biological Characteristics of Osteoblasts Derived from Myelodysplastic Syndrome Patients.

It was hypothesized that osteoblasts play a central role in hematopoiesis, and it has been shown that osteoblasts produce many factors essential for the survival, renewal, and maturation of hematopoietic stem cells (HSCs). By using human fetal osteoblastic cell line hFOB1.19 as a model of control, we investigated the biological characteristics of osteoblasts derived from patients with myelodysplastic syndrome (MDS) and their hematopoietic supportive function in vitro. MSCs isolated from bone marrow of MDS patients and normal donors were cultured and checked for their morphology, immunophenotype, CFU-F forming capacity and the expression of hematopoietic cytokines. A feeder layer was prepared by osteoblasts induced from 3rd generation of cultured MSCs and treated with mitomycin C. Ficoll-isolated bone marrow mononuclear cell from normal donors were then seeded on the feeder layer to co-culture in vitro without exogenous cytokines. FCM revealed that both MSCs and hFOB cells were positive for CD44, CD73(SH3), CD105(SH2) and CD90 (Thy1), but negative for CD34, CD45, HLA-DR. RT-PCR found that hFOB cells expressed mRNA of SCF, IL-6, IL-11, SDF-1, GM-CSF and G-CSF. MSCs obtained from MDS patients and normal donors were displaying fibroblastoid morphology. Their growth pattern, immunophenotype and CFU-F forming capacity were similar (P >0.05). Without exogenous cytokines, the osteoblasts derived from MDS could sustain GM-CFC survival for at least 3 weeks. The CFU-GM yield from cells in upper layer of co-culture was not different from those of control in hematopoiesis supportive experiments in vitro (P>0.05). RT-PCR clearly demonstrated that the cultured BM-MSCs from normal donor expressed mRNA of SCF, SDF-1, IL-6, and IL-11. As the MSCs differentiated toward osteoblasts, the expression of G-CSF could be detected, whereas GM-CSF remained undetectable. The same expression profile of above cytokines were also seen in osteoblasts induced from BM-MSCs of MDS patients. In conclusion, osteoblasts may play a pivotal role in hematopoiesis. The biological characteristics of osteoblasts from bone marrow of MDS patients were generally not different from those of osteoblasts in bone marrow of normal controls. Both of them could support survival of GM-CFC hematopoietic progenitor cells in vitro, according to their expression of multiple cytokines. These findings suggested that the osteoblasts derived from MDS patients may not be involved in the malignant process.

Influence of substratum surface chemistry/energy and topography on the human fetal osteoblastic cell line hFOB 1.19: Phenotypic and genotypic responses observed in vitro

Time-dependent phenotypic response of a model osteoblast cell line (hFOB 1.19, ATCC, and CRL-11372) to substrata with varying surface chemistry and topography is reviewed within the context of extant cell-adhesion theory. Cell-attachment and proliferation kinetics are compared using morphology as a leading indicator of cell phenotype. Expression of ( α 2, α 3, α 4, α 5, α v, β 1, and β 3) integrins, vinculin, as well as secretion of osteopontin (OP) and type I collagen (Col I) supplement this visual assessment of hFOB growth. It is concluded that significant cell-adhesion events—contact, attachment, spreading, and proliferation—are similar on all surfaces, independent of substratum surface chemistry/energy. However, this sequence of events is significantly delayed and attenuated on hydrophobic (poorly water-wettable) surfaces exhibiting characteristically low-attachment efficiency and long induction periods before cells engage in an exponential-growth phase. Results suggest that a ‘time–cell–substratum–compatibility–superposition principle’ is at work wherein similar bioadhesive outcomes can be ultimately achieved on all surface types with varying hydrophilicity, but the time required to arrive at this outcome increases with decreasing cell–substratum–compatibility. Genomic and proteomic tools offer unprecedented opportunity to directly measure changes in the cellular machinery that lead to observed cell responses to different materials. But for the purpose of measuring structure–property relationships that can guide biomaterial development, genomic/proteomic tools should be applied early in the adhesion/spreading process before cells have an opportunity to significantly remodel the cell–substratum interface, effectively erasing cause and effect relationships between cell–substratum–compatibility and substratum properties. Impact Statement This review quantifies relationships among cell phenotype, substratum surface chemistry/energy, topography, and cell–substratum contact time for the model osteoblast cell line hFOB 1.19, revealing that genomic/proteomic tools are most useful in the pursuit of understanding cell adhesion if applied early in the adhesion/spreading process.

Nanostructural characteristics, mechanical properties, and osteoblast response of spark plasma sintered hydroxyapatite

This study aimed to fabricate bulk nanostructured hydroxyapatite (HA) pellets with improved properties using spark plasma sintering (SPS) for orthopedic applications. Spray-dried nanostructured HA (nSD-HA) powders were consolidated using the rapid SPS processing. The SPS processed nSD-HA was characterized using Raman spectroscopy and field emission scanning electron microscopy (FESEM). Mechanical properties of the consolidates were also evaluated through indentation approach. The nanostructures ( approximately 80 nm in grain size) of the starting powders were successfully retained after the SPS processing operated at 950 degrees C with <15 min holding time. The SPS consolidated nSD-HA showed promising mechanical properties, approximately 118 GPa for Young's modulus, and up to 2.22 MPa m(0.5) for fracture toughness. SPS holding time showed minor influence on the phases of the pellets. Furthermore, the spheroidized nanostructured HA retained the HA structure after the SPS consolidation. Preliminary cytotoxicity and cell attachment studies were also carried out using a human osteoblast cell line hFOB 1.19. Enhanced cell attachment and proliferation on the nanostructured pellets were revealed. The presence of the nanostructures accounts mainly for the enhanced mechanical properties and promoted proliferation of the osteoblast cells. This study suggests that the SPS technique is an appropriate process for fabrication of bulk nSD-HA from nanostructured powder.

Culture media conditioned by heat‐shocked osteoblasts enhances the osteogenesis of bone marrow‐derived mesenchymal stromal cells

Osteogenic cells differentiated from bone marrow-derived mesenchymal stromal cells (MSC) hold much promise in bone tissue engineering and reconstructive surgery. There is a dire need for well-defined and efficient protocols to promote the osteogenesis of ex vivo cultured MSC. Hence, this study investigated whether a combination of chemical stimuli (ascorbic acid, beta-glycerophosphate and dexamethasone) and culture media conditioned by a human foetal osteoblast cell line (hFOB) had any synergistic effect on the osteogenesis of MSC. Conditioned media with or without prior heat shock treatment (42 degrees C for 1 h) of the hFOB cell line, were collected and tested on rabbit MSC cultures, in the presence and absence of chemical stimuli. Osteogenic differentiation of MSC was assessed on both day 14 and 21 of ex vivo culture. The results showed conclusively that conditioned media promoted osteogenesis of MSC, which was further enhanced by prior heat shock-treatment of the hFOB cells, as well as by the presence of chemical stimuli. Among all experimental groups, the combination of culture medium conditioned by heat shocked hFOB cells together with chemical stimuli, exhibited the highest level of calcium mineralization, as assessed by Von Kossa staining. This provides clear evidence of a synergistic effect of conditioned media, heat shock and chemical stimuli. It is hoped that the data may contribute to the development of a more well-defined and efficient in vitro culture protocol to promote the osteogenesis of MSC for both clinical and non-clinical applications.

Secretory products of breast cancer cells upregulate hyaluronan production in a human osteoblast cell line

Hyaluronan (HA) has been implicated in breast cancer progression and metastasis to lymph nodes. Although breast cancer has a strong propensity for metastasis to the bone, the role of HA in the development of bone metastasis in breast cancer is not well delineated. In order to determine the role of HA in breast cancer induced osteolysis, we examined the effect of secretory products in the conditioned medium of breast cancer cell lines on the expression of hyaluronan synthases (HAS), accumulation of HA in pericellular matrix, secretion of HA in the culture media, and on expression of surface HA receptors, in a human immature osteoblast cell line (Hfob). Our results show that conditioned medium derived from breast cancer cells upregulate the expression of hyaluronan synthases, HAS1 and HAS2, followed by significant increase in pericellular and secreted HA in Hfob cells. Our results further demonstrate that both CD44 and receptor for hyaluronan-mediated motility (RHAMM) are involved in binding cell surface associated HA on Hfob cells. Analysis of the growth factors in the conditioned medium implicates TGF-beta1 in the modulation of HAS1 and HAS2, as well as in the increase in pericellular and secreted HA. This report is the first to show that soluble factors produced by breast cancer cells mediate increase in HA production in osteoblasts.